Amoxycillin/clavulanate resistance as related to β-lactamase content in Escherichia coli strains from community-acquired urinary tract infections in Greece

Objective: To determine the resistance rate to amoxycillin/clavulanate (AMC) in 100 Escherichia coli strains isolated from outpatients with urinary tract infection (UTI) in four Greek hospitals and assess the relationship between β-lactamase content and resistance to AMC.
Methods: Susceptibility to β-lactams was determined with the E-test. Sonic cell extracts were used as β-lactamase preparations. Conjugal transfer of resistance was performed in broth cultures. β-Lactamase quantities were evaluated by measuring nitrocefin hydrolysis. Isoelectric points (pls) of β-lactamases were determined by electrofocusing. The substrate specificity of the enzymes and the inhibitory activity of clavulanate were studied spectrophotometrically.
Results: Thirty-two isolates were resistant to ampicillin. Eight were resistant (MIC ≥ 32 mg/L) and 11 showed decreased susceptibility (MIC 4–16 mg/L) to AMC. The latter expressed at least four-fold higher amounts of TEM-1 β-lactamase compared with the TEM-1-producing AMC-susceptible isolates. Seven AMC-resistant isolates produced at least 16-fold higher amounts of TEM-1; in one isolate, resistance was attributed to an OXA-type β-lactamase. None of the AMC-resistant isolates was able to transfer resistance to AMC by conjugation. Clavulanate-resistant TEM variants were not detected.
Conclusions: Amoxycillin/clavulanate-resistant E. coli strains have become established in the Greek community. Resistance is mainly due to the production of large amounts of TEM-1 β-lactamase which is encoded from non-self-transmissible plasmids.     

Resistance to β-lactams among blood isolates of Salmonella spp. in European hospitals: results from the SENTRY Antimicrobial Surveillance Program 1997–98

The susceptibility to β -lactams and the β -lactamase content of 110 Salmonella spp. blood isolates collected during 1997–98 in 19 European centers participating in the SENTRY Surveillance Program were studied. Thirty-one isolates (28%) were resistant to penicillins, due to production of TEM-1 (27 isolates), OXA-1 (three isolates) or TEM-1 + OXA-1 (one isolate). All OXA-1 producers and 10 TEM-1-producing isolates were also resistant to penicillin–clavulanic acid combinations. In the latter isolates, this phenotype was associated with increased production of TEM-1. Sixteen TEM-1-producing Salmonella Enteritidis isolates and one OXA-1-producing S. Typhimurium isolate were able to transfer β -lactam resistance by conjugation.     

Transferable plasmid mediating resistance to multiple antimicrobial agents in Klebsiella pneumoniae isolates in Greece

Objective   To investigate the underlying resistance mechanisms in 10 Klebsiella pneumoniae isolates.
Methods   Ten K. pneumoniae strains according to distinct bacteriocin typing and REP-PCR, were examined for their plasmid content, their ability to transfer their resistance to aminoglycosides and third-generation cephalosporins, and their production of aminoglycoside-modifying enzymes and β -lactamases.
Results   Transfer of resistance to the above-mentioned antibiotics as well as to co-trimoxazole and tetracycline in Escherichia coli strain RC 85 at a frequency of 5–10⁶ was achieved for all strains by conjugation. Similar strains harbor a self-transferable multiresistant plasmid (80 kb) with similar Eco RI and Hind III restriction patterns. This plasmid encodes an extended-spectrum β -lactamase which confers high-level resistance to third-generation cephalosporins and aztreonam. It produces SHV-5 β -lactamase, as demonstrated by isoelectric focusing and DNA sequencing. Aminoglycoside resistance was co-transferred, and AAC(6')-I, mediating resistance to gentamicin, tobramycin, netilmicin and amikacin, and AAC(3)-I, mediating resistance to gentamicin and sisomycin, were encoded in all isolates and their transconjugants, while APH(3')-I, mediating resistance to kanamycin and neomycin, was encoded in seven strains.
Conclusions   It appears that a multiresistant transferable plasmid encoding the SHV-5 β -lactamase, causing unusually high resistance to ceftazidime and aztreonam, and the combination AAC(6')-I + AAC(3)-I of acetylating enzymes causing, also resistance to all clinically available aminoglycosides, is established in K. pneumoniae in Greece.     

Antibiotic susceptibilities of Haemophilus influenzae in central Scotland

Objective: To ascertain the incidence of antibiotic resistance in Haemophilus influenzae in central Scotland and the β-lactamases produced by these isolates.
Methods: A total of 213 H. influenzae isolates from four medical centers in Scotland [Aberdeen ( n = 58), Edinburgh ( n = 55), Glasgow ( n = 64) and Dundee ( n = 36)] were tested for susceptibility to a range of antimicrobials including β-lactams, β-lactam/β-lactamase-inhibitor combinations, and a representative 4-quinolone, antifolate and macrolide. Susceptibility testing of the β-lactam/β-lactamase-inhibitor combination amoxicillin plus clavulanic acid was conducted at both 2:1 and 4:1 ratios and with clavulanic acid fixed at a concentration of 2 mg/L. Each strain was further investigated for the presence of β-lactamase activity.
Results: Although the incidence of resistance to amoxicillin was 15%, in the presence of clavulanic acid, this resistance was reduced to 4.2%, 5.6% and 4.2% with the 2:1 ratio, 4:1 ratio and 2 mg/L fixed concentration, respectively. Sixteen percent of the isolates demonstrated immediate β-lactamase production. Isoelectric focusing showed that 77.4%, 16.1% and 6.5% of the β-lactamase-positive strains were found to contain TEM-1, VAT-1 and both TEM-1 and VAT-1 β-lactamases, respectively. A further 29% of the strains were recognized as being β-lactamase-positive after prolonged incubation with nitrocephin.
Conclusions: This study suggests that current testing for β-lactamases may underestimate the prevalence of β-lactamase production in H. influenzae.     

Incidence, epidemiology and evolution of reduced susceptibility to ciprofloxacin in Neisseria gonorrhoeae in Korea

Objective: To verify the decrease of susceptibility to ciprofloxacin in Neisseria gonorrhoeae , determine the size of the recently reported new β-lactamase plasmid and explain the high prevalence of penicillinase-producing Neisseria gonorrhoeae (PPNG).
Methods: Gonococci were isolated from prostitutes in Korea. Antimicrobial susceptibility was tested by NCCLS disk diffusion and agar dilution methods. Plasmid was isolated by an alkaline lysis method. Patterns of Nhe l-digested genomic DNA were compared after pulsed-field gel electrophoresis (PFGE).
Results: The minimum inhibitory concentration of ciprofloxacin for 50% of the isolates rose from 0.015 mg/L in 1993 to 0.12 mg/L in 1996. The proportion of PPNG remained at 70% or over during the 5-year period. The size of a novel β-lactamase plasmid, first reported in 1994, was determined to be approximately 3.2 MDa, and 48% of the PPNG isolates contained it. Twelve of 50 isolates had the same PFGE pattern and nine others another pattern.
Conclusion: The rapid decrease of fluoroquinolone-susceptible gonococci suggests that in the near future the drug may become less useful for gonorrhea treatment. The new 3.2-MDa plasmid may have been introduced as a result of the recent increase in overseas travel. The PFGE pattern suggests that high prevalence of PPNG may be due to dissemination of a few resistant clones among the high-risk groups.     

Antibiotic susceptibility and genotypic characterization of Haemophilus influenzae strains isolated from nasopharyngeal specimens from children in day-care centers in eastern France

Objective   To determine the overall carriage rate for Haemophilus influenzae in young children in day-care centers, the frequency of resistance to various classes of antibiotic, and the clonal relationship between isolates of the various resistant phenotypes.
Methods   Nasopharyngeal (NP) specimens were obtained and cultured on chocolate agar with bacitracin. Antibiotic susceptibility testing and serotyping were performed for all isolates. The genetic polymorphism of ampicillin-susceptible and β-lactamase-producing isolates was studied by pulsed-field gel electrophoresis using Sma I.
Results   Of the 596 NP secretion cultures, 152 (25.5%) were positive for H. influenzae . Sixty-four (42.1%) isolates produced β-lactamase and two (1.3%) were ampicillin resistant but did not produce β-lactamase. We were unable to serotype 150 isolates; one isolate belonged to capsular serotype e and one to serotype f. Forty-six major DNA patterns were identified among 76 randomized isolates. β-lactamase producing isolates more frequently showed EP than ampicillin-susceptible isolates P  < 10⁻⁴. The frequency of isolates with EP was significantly lower in day-care centers attended by less than 20 children than in those attended by more than 20 children ( P  = 0.020).
Conclusions   Resistance due to β-lactamase production has disseminated in some day-care centers, mostly by person-to-person spread but also via the possible conjugal transfer of large plasmids between strains. The size of day-care centers may affect the risk of transmission.     

Bactericidal activity of three β-lactams alone or in combination with a β-lactamase inhibitor and two aminoglycosides against Klebsiella pneumoniae harboring extended-spectrum β-lactamases

Objective: To study the bactericidal activity of β-lactam antibiotics (imipenem, cefepime, cefpirome) alone or in combination with a β-lactamase inhibitor (sulbactam) in the presence or absence of aminoglycoside (amikacin or isepamicin) against Klebsiella pneumoniae strains producing extended-spectrum β-lactamases (ESBLs).
Methods: We characterized 10 strains by means of analytic isoelectric focusing and pulsed-field gel electrophoresis. The ESBLs produced by these strains were derived from either TEM (TEM-1, TEM-2) or SHV-1. The killing-curve method was used for this bacterial investigation. Bacteria (final inoculum 5×10 ⁵ CFU/mL) were incubated with antibiotics at clinical concentrations obtained in vivo.
Results: All the combinations with cefepime or cefpirome + sulbactam were bactericidal, with a 4 log₁₀ decrease being obtained within 6 h without regrowth at 24 h, whereas imipenem alone, and combinations, gave a bactericidal effect within 6 h. The two cephalosporins alone decreased the inoculum of 4 log₁₀ at 6 h but regrowth was observed at 24 h. When the aminoglycoside was added, this bactericidal effect was obtained within 3 h with amikacin and within 1 h with isepamicin.
Conclusions: Cefepime + sulbactam or cefpirome + sulbactarn may be an alternative to imipenem for the treatment of patients with ESBL-producing K. pneumoniae. Aminoglycosides are often associated in nosocomial infections due to ESBL-producing K. pneumoniae: isepamicin acted faster than amikacin, but both worked well. To conclude, it may be prudent to avoid extended-spectrum cephalosporins as single agent when treating serious infections due to ESBL-producing K. pneumoniae. Addition of a β-lactamase inhibitor such as sulbactam ± aminoglycoside is advisable to avoid failure of treatment.     

Emergence and outbreaks of CTX-M beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae strains in a Tunisian hospital

Sixty-two isolates of Enterobacteriaceae (35 Escherichia coli and 27 Klebsiella pneumoniae isolates) producing CTX-M-type beta-lactamases were collected between March 2000 and June 2003 in different wards of Charles Nicolle Hospital in Tunis (Tunisia). Sequencing identified the bla(CTX-M-15) determinant in 55 isolates and bla(CTX-M-16) in 7 isolates. The CTX-M-15-producing strains were isolated in several wards and consisted mainly of two successive clonal groups of E. coli and a major clonal group of K. pneumoniae. The second clonal group of E. coli belonged to phylogenetic group B2 and harbored more virulence factors than the first clonal group. Among the 22 transconjugants or electroporants obtained with selected E. coli and K. pneumoniae CTX-M-15-producing strains, a predominant plasmid restriction pattern was obtained with 17 isolates. The four CTX-M-16-producing strains of E. coli yielded the same pulsed-field gel electrophoresis (PFGE) pattern, while the three CTX-M-16-producing strains of K. pneumoniae yielded two different PFGE patterns. All of the CTX-M-16-producing isolates were recovered in the pediatric ward and had the same plasmid restriction pattern.     

Metallo-β-lactamase-producing Pseudomonas aeruginosa isolated from a large tertiary centre in Kenya

This study was designed to characterize the β-lactamase content of carbapenem-resistant Pseudomonas aeruginosa isolates recovered during 2006 and 2007 in a large tertiary-care centre in Nairobi, Kenya. Molecular characterization was done using PCR and sequencing, and typing was performed using pulsed-field gel electrophoresis (PFGE). In total, 416 P. aeruginosa isolates were obtained during that period, of which 57 (13.7%) were resistant to carbapenems. All carbapenem-resistant isolates tested positive for metallo-β-lactamase (MBL) production. All MBL isolates produced VIM-2 with two types of integron structures . PFGE identified three clonally related groups of VIM-2-producing P. aeruginosa , including a pan-resistant clone that was responsible for nosocomial outbreaks during 2006 and 2007 in the intensive-care unit. These findings suggest that continuous molecular surveillance needs to be performed to monitor the spread within the hospital of this pan-resistant strain. This study is the first report of VIM-2-producing P. aeruginosa from the African continent.     

Bacterial and epidemiologic study of the resistance to oxyiminocephalosporins in Escherichia coli in a French hospital

Objective: To understand the mechanisms and epidemiology of resistance to oxyiminocephalosporins in Escherichia coli over a 2-year period in a French hospital.
Methods: Forty-four strains, resistant or intermediately resistant to one of the oxyiminocephalosporins or aztreonam, were collected from 35 patients. MIC determinations were carried out for the 44 isolates using a panel of β-lactam antibiotics, and characterization of the β-lactamases they produced by isoelectric focusing and catalytic activity measurement. Extended-spectrum β-lactamase production was studied by use of the double disk diffusion test. Conjugation experiments were used to search for plasmidic cephalosporinase. An epidemiologic study was then performed, by use of molecular typing of the strains with an ERIC-PCR method and a case-control analysis.
Results: Less than 1% of all the E. coli isolates at our hospital showed decreased susceptibility to oxyiminocephalosporins. Only three of the 44 isolates showed synergy between clavulanate and a third-generation cephalosporin and produced an extended-spectrum β-lactamase. For the other strains, a β-lactamase with a highly basic isoelectric point was detected. Spectrophotometric measures confirmed that most of these isolates were AmpC hyper-producers. No plasmidic cephalosporinase could be detected by conjugation experiments. Molecular typing showed all isolates to be different, except for two strains isolated in two patients of the same hospital unit, and for the repeated isolates of some patients. When 20 case patients were compared to 40 randomly selected control patients, prior receipt of an antimicrobial and more specifically of a β-lactam agent was significantly associated with case patients.
Conclusions: Although it appears to be very rare, the resistance to broad-spectrum cephalosporins needs our attention, because of the high frequency of E. coli infections and β-lactam use in their treatment.     

Molecular analysis of ampicillin-resistant sporadic Salmonella typhi and Salmonella paratyphi B clinical isolates

Objective: To study the mechanisms of antibiotic resistance in Salmonella typhi and Salmonella paratyphi B clinical isolates, and the clonality of resistant strains.
Method: Antibiotic susceptibility was tested by disk-agar diffusion. Conjugation experiments and plasmid analysis by agarose gel electrophoresis after Eco RI digestion were followed by hybridization to a digoxigenin-labeled TEM-type β-lactamase probe. DNA fingerprints were obtained by pulsed-field gel electrophoresis of Xba I-digested chromosomal DNA.
Results: Three S. typhi isolates (7% of the isolates studied), of which one was ampicillin resistant and the other two multiresistant (ampicillin, chloramphenicol, tetracycline, sulfamethoxazole/trimethoprim and streptomycin), and two ampicillin-resistant S. paratyphi B isolates (25% of the isolates studied) were further evaluated. A 34-MDa conjugative plasmid, previously isolated from Salmonella enteritidis , conferred ampicillin resistance. A 100-MDa conjugative plasmid encoded resistance to chloramphenicol, tetracycline and sulfamethoxazole/trimethoprim, as well as ampicillin. Chromosomal fingerprinting revealed two distinct resistant strains for each serovar which were different from a matched set of sensitive S. typhi strains.
Conclusions: Two conjugative, TEM-type β-lactamase-encoding plasmids conferred ampicillin resistance to S. typhi and S. paratyphi B. The 34-MDa plasmid was identical to that previously characterized from S. enteritidis , while the 100-MDa plasmid also encoded resistance to chloramphenicol, tetracycline and sulfamethoxazole/trimethoprim. Resistant isolates did not belong to a single clone but rather represented distinct strains.     

Epidemiology of extended-spectrum beta-lactamase-producing Enterobacter isolates in a Spanish hospital during a 12-year period

Fifteen Enterobacter clinical isolates (11 Enterobacter cloacae isolates, 3 Enterobacter aerogenes isolates, and 1 Enterobacter gergoviae isolate), representing 0.4% of all Enterobacter isolates recovered in our hospital from 1989 to 2000, were suspected of harboring an extended-spectrum beta-lactamase (ESBL). These isolates were recovered from 14 different patients. ESBLs were transferred by conjugation into an Escherichia coli recipient strain. Pulsed-field gel electrophoresis (PFGE) revealed a single clone of E. aerogenes and six different clones of E. cloacae. Four of these E. cloacae clonal types were represented by only one isolate each, but the other two were represented by three and four isolates, respectively. Isoelectric focusing, susceptibility phenotyping, PCR analysis, and sequencing demonstrated the presence of three different ESBLs. The most frequent was the recently characterized CTX-M-10 ESBL, which was found in the E. gergoviae isolate and in all but one of the E. cloacae isolates. The remaining E. cloacae isolate harbored a TEM-27 ESBL, and the three E. aerogenes isolates harbored a TEM-24 ESBL. PFGE revealed that our E. aerogenes strain was indistinguishable from the French TEM-24-producing E. aerogenes endemic clone. Although a low prevalence of ESBL-producing Enterobacter isolates was found in our institution over a 12-year period, a diversity of nonepidemic E. cloacae clones was detected, as was the persistence of the CTX-M-10 beta-lactamase. The presence of the TEM-24-producing E. aerogenes French clone in our institution also demonstrates the intercountry dissemination of ESBL-producing isolates.     

Evaluation of clonal relatedness of extended-spectrum β-lactamase-producing Proteus mirabilis isolates by quantitative antibiogram and RAPD typing

Objective: To delineate, using two different typing systems, the clonal relatedness of 40 isolates of extended-spectrum β-lactamase (ESβla)-producing Proteus mirabilis obtained over a period of 7 years in six hospitals in the Paris area and two in Pas-de-Calais.
Method: Random amplified polymorphic DNA (RAPD) polymerase chain reaction typing was applied by using three random primers on the ESβla-producing P. mirabilis isolates and on isogenic Escherichia coli strains with or without plasmids encoding the representative resistance pattern transferred from P. mirabilis. Quantitative antibiogram typing, which was also applied to the P. mirabilis isolates, was used to define the euclidean distance between these strains.
Results: After having demonstrated that P. mirabilis plasmids did not influence chromosomal DNA amplification, we could classify the ESβla-producing P. mirabilis isolates into 12 groups based on RAPD fingerprints. The same isolates were classified into 19 groups by quantitative antibiogram typing. Despite this difference in group numbers, general concordance between the typing systems was observed. This allowed us to show that the greater number of isolates in some hospitals belonged to a single strain and that single isolates obtained in different hospitals generally represented unique strains.
Conclusions: A small number of ESβla-producing P. mirabilis strains was isolated during 7 years in the eight medical centers studied, and the number of different strains identified suggested that inter-hospital transfer had not occurred.     

Phage types, antibiotic susceptibilities and plasmid profiles of Salmonella typhimurium and Salmonella enteritidis strains isolated in Istanbul, Turkey

Objective   To examine 13 Salmonella typhimurium and 22 S. enteritidis strains isolated from individual cases of gastroenteritis for their phage types, antibiotic susceptibilities and plasmid profiles.
Methods   The phage typing of S. typhimurium strains was done according to the method of Anderson et al, and the phage typing scheme of Ward et al was used for phage typing of S. enteritidis strains. Antibiotic susceptibility testing was performed by the Kirby–Bauer disk diffusion method. Extended-spectrum β-lactamase production of the strains was determined by the three-dimensional method. Plasmid profiles of the strains were examined using the method described by Kado and Liu with some modification by Graeber et al.
Results   Two S. typhimurium strains were DT 193 and one was DT 22, whereas 10 strains were untypable. PT 4 was the predominant phage type among S. enteritidis strains. Four S. enteritidis strains were DT 6a, three strains were PT 1 and one strain was PT 8, whereas only one strain was untypable. Eleven of 13 S. typhimurium and three of 22 S. enteritidis strains were found to be multiresistant. Ten different resistance patterns among S. typhimurium and four different resistance patterns among S. enteritidis strains were detected. Extended-spectrum β-lactamase production was detected in 10 of 13 S. typhimurium and in three of 22 S. enteritidis strains. All S. typhimurium strains but one were found to contain at least one plasmid, with molecular masses varying between 4 and 107 MDa, and 11 different plasmid patterns were determined. Plasmid pattern analysis permitted further differentiation of the S. enteritidis strains into nine groups. A serovar-specific virulence plasmid of 36 MDa was detected in 13 of 22 S. enteritidis strains.
Conclusions   The results suggest that the majority of S. typhimurium strains were closely related.     

Carbapenem-hydrolyzing class D β-lactamase OXA-48 in Klebsiella pneumoniae isolates from Tunisia

Two carbapenem-resistant Klebsiella pneumoniae isolates producing the plasmid-encoded carbapenem-hydrolyzing OXA-48 were identified. These isolates, recovered from two patients hospitalized in two different hospitals in Tunisia in December 2010, were not clonally related. Molecular investigations showed that both isolates co-produced the narrow-spectrum β-lactamases TEM-1 and SHV-1, together with the extended-spectrum β-lactamase CTX-—15.     

Genotyping,local prevalence and international dissemination of β-lactamase-producing Kingella kingae strains

β-lactamase production has been sporadically reported in the emerging Kingella kingae pathogen but the phenomenon has not been studied in-depth. We investigated the prevalence of β-lactamase production among K. kingae isolates from different geographical origins and genetically characterized β-lactamase-producing strains. Seven hundred and seventy-eight isolates from Iceland, the USA, France, Israel, Spain and Canada were screened for β-lactamase production and, if positive, were characterized by PFGE and MLST genotyping, as well as rtxA, por, bla_TEM and 16S rRNA sequencing. β-lactamase was identified in invasive strains from Iceland (n = 4/14, 28.6%), the USA (n = 3/15, 20.0%) and Israel (n = 2/190, 1.1%) and in carriage strains in the USA (n = 5/17, 29.4%) and Israel (n = 66/429, 15.4%). No French, Spanish or Canadian isolates were β-lactamase producers. Among β-lactamase producers, a perfect congruency between the different typing methods was observed. Surprisingly, all US and Icelandic β-lactamase-producing isolates were almost indistinguishable, belonged to the major international invasive PFGE clone K/MLST ST-6, but differed from the four genetically unrelated Israeli β-lactamase-producing clones. Representative strains of different genotypes produced the TEM-1 enzyme. K. kingae β-lactamase producers exhibit a clear clonal distribution and have dissimilar invasive potential. The presence of the enzyme in isolates belonging to the major worldwide invasive clone K/ST-6 highlights the possible spread of β-lactam resistance, and emphasizes the importance of routine testing of all K. kingae clinical isolates for β-lactamase production.     

Molecular Epidemiology of Extended-Spectrum β-Lactamase-Producing Klebsiella pneumoniae Isolates in a District Hospital in Taiwan

Thirty-one of 104 clinical isolates of Klebsiella pneumoniae collected over a period of 8 months were found to be putative extended-spectrum β-lactamase (ESBL) producers. Isoelectric focusing and an iodine overlay agar method were used for preliminary identification of the ESBLs. They were further identified by DNA sequencing. Seventy-one percent of the isolates were found to produce SHV-5. The variation in the ESBL patterns of these isolates was slight, with only five patterns being identified. The strains were typed by pulsed-field gel electrophoresis (PFGE), and 16 different genotypes were identified. When the PFGE patterns were analyzed by the algorithmic clustering method called the unweighted-pair group method using arithmetic averages, five clusters were found. However, significant genetic variations were found among 11 isolates and between each cluster. A plasmid of 36 kb was found in all clinical isolates and in the transconjugants. Our results indicate that the increase in the number of ESBL-producing K. pneumoniae isolates in this hospital is due mainly to the dissemination of a resistance plasmid rather than to the clonal spread of a few epidemic strains.     

Antibiotic Resistance Patterns and Plasmid Profiles of Salmonella typhimurium Isolates in Turkey

 To understand the resistance mechanisms present in 75 isolates of Salmonella typhimurium derived from clinical infections in Turkey, antimicrobial resistance patterns and associated plasmids were investigated. Among the 22 strains that produced extended-spectrum β-lactamase (ESBL), 20 were resistant to aminoglycosides and 12 to trimethoprim-sulfamethoxazole. Strains that did not produce ESBL did not express aminoglycoside or trimethoprim-sulfamethoxazole resistance, although 27 of them were ampicillin resistant. None of the strains were resistant to imipenem or fluoroquinolones. Nineteen strains producing ESBL carried a plasmid of >100 MDa. Seven ESBL-producing strains conjugally transferred their ESBLs and trimethoprim-sulfamethoxazole resistance. No correlation was found between the resistance patterns and plasmids in non-ESBL-producing strains.     

Cefotaxime-Hydrolysing Beta Lactamases in Morganella morganii

 The frequency of enterobacterial isolates with high resistance to expanded-spectrum β-lactam antibiotics (mainly cefotaxime or ceftriaxone) has increased notoriously in Argentina, mainly because of the spread of extended-spectrum β-lactamases. The aim of this work was the study of extended-spectrum β-lactamases in several Morganella morganii isolates with unusually high resistance to ceftriaxone. These strains produced at least two β-lactamases, of apparent pIs of 5.4 and 8.2, molecular weight 23 000, well inhibited by clavulanate, compatible with a broad-spectrum β-lactamase – perhaps TEM-1 – and an extended-spectrum β-lactamase, respectively. The extended-spectrum β-lactamase was identified as a CTX-M-type β-lactamase – probably CTX-M-2 – by polymerase chain reaction, restriction profile analysis and DNA-DNA hybridisation. The remaining isolates studied produced either the broad-spectrum β-lactamase plus the ubiquitous AmpC β-lactamase (13 strains), or the AmpC β-lactamase only (10 strains).     

Evaluation of the molecular epidemiology of an outbreak of multiply resistant Shigella sonnei in a day-care center by using pulsed-field gel electrophoresis and plasmid DNA analysis.

Outbreaks of diarrhea in child day-care centers (DCC) are common. This study was undertaken to evaluate the molecular epidemiology of an outbreak of diarrhea due to Shigella sonnei. This outbreak involved 25 of 52 (48%) DCC children and 14 of 132 (11%) teachers and household contacts. S. sonnei isolates from nine children and five contacts were characterized by antimicrobial susceptibility, plasmid content, plasmid DNA restriction fragment pattern, and pulsed-field gel electrophoresis (PFGE) of total genomic DNA; 33 isolates from Houston, Tex., Chicago, Ill., and Mexico City, Mexico, also were studied. All outbreak isolates were resistant to ampicillin and trimethoprim-sulfamethoxazole and shared five to six plasmids ranging from 3.3 to 70 MDa. A total of 8 of 12 temporally associated nonoutbreak Houston isolates had plasmid profiles and restriction fragment patterns similar to those of the outbreak strain, despite possessing different antibiotic susceptibility patterns. PFGE demonstrated identical DNA patterns among outbreak isolates and similar or identical patterns among temporally associated sporadic Houston isolates with plasmid profiles similar to that of the outbreak strain. All other nonoutbreak strains from Houston, Chicago, and Mexico had plasmid profiles, restriction fragment patterns, and PFGE patterns different from those of the outbreak strain. DCC outbreak isolates could be distinguished from most sporadic isolates by antimicrobial susceptibility testing, but plasmid analysis and PFGE could not differentiate common-source isolates from sporadic isolates in the same location during the same time period, indicating that isolates present in the community were genetically similar to those producing outbreaks in the DCC.