Antibiotic-resistant Klebsiella Research Articles

Suppression of stop codon UGA in acrB can contribute to antibiotic resistance in Klebsiella pneumoniae ATCC10031

We previously reported that Klebsiella pneumoniae MGH78578 exhibited higher resistance against various antimicrobials than K. pneumoniae ATCC10031. In this study, we showed that the plasmid, pKPN5, in K. pneumoniae MGH78578 played an important role in resistance against aminoglycosides, ampicillin, tetracycline, and chloramphenicol, while genome-derived β-lactamases and drug efflux pumps appeared to be more important in resistance to cloxacillin. acrAB, encoding a potent multidrug efflux pump, was cloned from K. pneumoniae MGH78578 and ATCC10031, to investigate reasons for the high drug resistance of K. pneumoniae MGH78578, and the results revealed that AcrAB from K. pneumoniae ATCC10031 conferred weaker drug resistance than AcrAB from K. pneumoniae MGH78578. DNA sequencing revealed that acrB from K. pneumoniae ATCC10031 carried the nonsense mutation, UGA, which was not found in acrB from K. pneumoniae MGH78578. However, acrB from K. pneumoniae ATCC10031 conferred slightly elevated resistant levels to several antimicrobials. The intact length of AcrB was detected in K. pneumoniae ATCC10031 by Western blot analysis, even though its quantity was small. Therefore, the stop codon UGA in acrB was thought to be overcome to some extent in this strain. We artificially introduced the nonsense mutation, UGA to the cat gene on pACYC184, and the plasmid also elevated the MIC of chloramphenicol in K. pneumoniae ATCC10031. These results suggest that a mechanism to overcome the nonsense mutation in acrB sustained resistance against a few β-lactams, dyes, and cholic acid in K. pneumoniae ATCC10031.

Influence of induced ciprofloxacin resistance on efflux pump activity of Klebsiella pneumoniae

The efflux pump (EP) is one of the major mechanisms of antibiotic resistance in Klebsiella pneumoniae. However, there are few reports on the effect of the abuse of antibiotic use on the activity of EPs. To determine whether the use of low efficacy antibiotics has any effect on the activity of EPs and induces drug resistance in K. pneumoniae, we investigated the effect of ciprofloxacin on the activity of EPs in K. pneumoniae strains. Sixteen susceptible K. pneumoniae strains were isolated from patients and their minimum inhibitory concentrations (MICs) of ciprofloxacin were measured in the absence and presence of the pump inhibitor carbonyl cyanide m-chlorophenyl hydrazone (CCCP). The strains were then induced with a gradient of ciprofloxacin until the MICs of the strains showed no further increase, to obtain induced resistant strains. The EP activities of the strains before and after induction were compared using EP inhibition and ethidium bromide (EtBr) accumulation assays. The MIC values of the strains were 16‒256 times higher after induction than before induction. In the presence of CCCP, the MIC values of 50% of the induced strains were 2‒4-fold lower than that in the absence of this inhibitor. The EtBr accumulation assay showed that the fluorescence of EtBr in the induced cells was lower than that in the cells before induction. EPs are widespread in susceptible and drug-resistant K. pneumoniae strains. Induction with ciprofloxacin may increase the activity of EPs in K. pneumoniae. The EtBr accumulation assay is more sensitive than the EP inhibition assay in evaluating the activity of EPs in K. pneumoniae.

Outcomes of pancreatic resection: Analysis of the french national database on 6436 patients

Outcomes of pancreatic resection: Analysis of the french national database on 6436 patients

Outbreak of antibiotic-resistant klebsiella pneumoniae in a high-volume pancreatic surgery unit

Outbreak of antibiotic-resistant klebsiella pneumoniae in a high-volume pancreatic surgery unit

Susceptibility rate to tigecycline and antibiotic-resistance among Klebsiella pneumoniae strains isolated in Intensive Care Unit

Antibiotic resistance in Klebsiella pneumoniae strains is an increasing problem in a lot of hospitals. It is a public health emergency because it relates with high mortality rate among patients in Intensive Care Unit (ICU). From 1/1/2009 to 31/08/2010, in ICU of SS Annunziata Hospital of Taranto, 140 isolated Klebsiella pneumoniae strains were detected. The strain identification and antimicrobial susceptibility testing were performed using a Vitek2 automated system. These isolate showed a low level of susceptibility to levofloxacin (3.4%), ciprofloxacin (6.2%), ceftazidime (2.8%) and piperacillin/tazobactam (8%). We reported also that the 10% and 13.9% of them were susceptible to meropenem and imipenem. An anti-Klebsiella pneumoniae activity in vitro to tigecycline was present in 64.6% of isolates while almost all strains (56/58) tested to colistin were susceptible. In order to our data of worryng high multiclass drug resistance including tygecicline, it needs to apply appropriate measures of surveillance and antibiotic prescription to avoid rapid spread of these mutiresistant strains in other areas.

Isolation and characterization of multidrug-resistant Klebsiella spp. isolated from shrimp imported from Thailand

A study was undertaken to isolate and characterize tetracycline and nalidixic acid-resistant Klebsiella spp. in farm-raised, imported shrimp sold in the United States. Sixty-seven multiple antibiotic-resistant Klebsiella spp. strains were isolated from imported shrimp samples. Using morphological and biochemical methods, fifty-three strains were tentatively identified as Klebsiella pneumoniae and fourteen as K. oxytoca. Although all isolates were resistant to tetracycline, only 8 were resistant to nalidixic acid. These 8 isolates were further screened by PCR for quinolone resistance genes (qnrA, B, S, gyrA, B and parC). PCR protocols failed to amplify any qnr genes. The purified PCR amplicons of gyrA, gyrB and parC were sequenced and analyzed for point mutations that confer resistance to fluoroquinolone antibiotics. Analysis of the sequences of the gyrA amplicons from nalidixic acid-resistant Klebsiella spp. indicated two point mutations in gyrA at positions 83 (Ser→Phe) and 87 (Asp→Ala). Sequence analysis of the parC amplicons indicated an amino acid change at position 80 (Ser→Ile). No mutations were detected in gyrB. Template DNA from all isolates was screened for tetracycline resistance genes (tetA–E). Oligonucleotide primers specifically targeting a 305-bp region of tetB and a 477-bp region of tetD successfully amplified sequences from 91.0 and 44.0% of the isolates, respectively. None of the isolates contained tetA, tetC or tetE genes. Plasmids (2.0–16.0kb) were found in 23 of the 67 isolates. XbaI-PFGE identified 32 distinct macro restriction patterns (mrps) among the 61 multiple drug-resistant Klebsiella spp. that were typable. Our results indicate that imported shrimp is a reservoir for multidrug resistant Klebsiella spp. and potential health risks posed by such strains should not be underestimated.

A potential antimicrobial treatment against ESBL-producing Klebsiella pneumoniae using the tellurium compound AS101

Due to the extensive spread of antibiotic-resistant Klebsiella pneumoniae, the non-toxic immunomodulator, ammonium trichloro (dioxoethylene-o, o') tellurate (AS101), was introduced for the first time in this study. Eleven strains of K. pneumoniae were tested: five were extended spectrum beta lactamase (ESBL)-producing strains and six were non-ESBL-producing strains. The MIC and MBC of ten strains were 9 microg/ml AS101 and 18 microg/ml for one strain. AS101 treatment inhibited bacterial growth in a dose-dependent manner on protein-rich media. No inhibition by AS101 was observed on poorer media. In combination with beta-mercaptoethanol (2-ME) or cysteamine, AS101 inhibited bacterial growth in both types of media. Growth inhibition was also shown following AS101 treatment at both lag and log phases. Our data indicate that AS101 enters the bacterium through its porins, causing bacterial destruction. The mechanism of cell death was characterized using several techniques: (a) scanning electron microscopy showed that bacteria treated with AS101 or in combination with cysteamine exhibited evidence of cell-wall damage; (b) X-ray microanalysis demonstrated damage to Na/K pumps; and (c) transmission electron microscopy demonstrated cell lysis. These phenomena suggest that AS101 has antibacterial potential against K. pneumoniae infections.

Epidemiology of an outbreak of antibiotic-resistant Klebsiella pneumoniae at a tertiary care medical center

In 2004, a 650-bed, tertiary care medical center experienced an outbreak of multiple antibiotic-resistant Klebsiella pneumoniae (MR-KP) that included extended-spectrum beta-lactamase (ESBL)-producing and non-ESBL-producing strains. Characteristics associated with MR-KP were evaluated by case-control study with variables tested by conditional regression analyses. Pulsed-field gel electrophoresis (PFGE) was used to compare the molecular relatedness of isolates. In 2004, the incidence rate of MR-KP increased significantly compared with 2003 (relative risk [RR], 5.1; 95% confidence interval [CI]: 3.10-8.37) when only ESBL-producing K pneumoniae were present. The increase involved both ESBL-producing MR-KP and MR-KP in which ESBL production was not detected by the testing in use. Nineteen isolates were identical or closely related by PFGE. Characteristics associated with MR-KP were longer length of hospital stay (odds ratio [OR], 2.92; 95% CI: 1.17-7.30; P = .022), greater total antibiotic-days (OR, 2.81; 95% CI: 1.19-6.65; P = .018], and higher Acute Physiology and Chronic Health Evaluation (APACHE) II score (OR, 1.15; 95% CI: 1.06-1.25; P = .001). When the MR-KP cases were subdivided into ESBL-producing K pneumoniae and ESBL-negative K pneumoniae, while controlling for length of stay, total antibiotic-days was significantly associated with ESBL-producing K pneumoniae (OR, 3.8; 95% CI: 1.2-12.1; P = .02). Compared with patients housed on the same unit at the same time, patients with MR-KP had a longer length of stay and greater antibiotic exposure. Patients with longer length of stay and greater total antibiotic exposure should be potential targets for stringent infection control measures.

Genome Sequencing and Comparative Analysis ofKlebsiella pneumoniaeNTUH-K2044, a Strain Causing Liver Abscess and Meningitis

Nosocomial infections caused by antibiotic-resistant Klebsiella pneumoniae are emerging as a major health problem worldwide, while community-acquired K. pneumoniae infections present with a range of diverse clinical pictures in different geographic areas. In particular, an invasive form of K. pneumoniae that causes liver abscesses was first observed in Asia and then was found worldwide. We are interested in how differences in gene content of the same species result in different diseases. Thus, we sequenced the whole genome of K. pneumoniae NTUH-K2044, which was isolated from a patient with liver abscess and meningitis, and analyzed differences compared to strain MGH 78578, which was isolated from a patient with pneumonia. Six major types of differences were found in gene clusters that included an integrative and conjugative element, clusters involved in citrate fermentation, lipopolysaccharide synthesis, and capsular polysaccharide synthesis, phage-related insertions, and a cluster containing fimbria-related genes. We also conducted comparative genomic hybridization with 15 K. pneumoniae isolates obtained from community-acquired or nosocomial infections using tiling probes for the NTUH-K2044 genome. Hierarchical clustering revealed three major groups of genomic insertion-deletion patterns that correlate with the strains' clinical features, antimicrobial susceptibilities, and virulence phenotypes with mice. Here we report the whole-genome sequence of K. pneumoniae NTUH-K2044 and describe evidence showing significant genomic diversity and sequence acquisition among K. pneumoniae pathogenic strains. Our findings support the hypothesis that these factors are responsible for the changes that have occurred in the disease profile over time.

Antibiotic-resistant Klebsiella pneumoniae in Spain: analyses of 718 invasive isolates from 35 hospitals and report of one outbreak causedby an SHV-12-producing strain

Antibiotic-resistant Klebsiella pneumoniae in Spain: analyses of 718 invasive isolates from 35 hospitals and report of one outbreak causedby an SHV-12-producing strain

Incidence of antibiotic-resistant Klebsiella pneumoniae and Enterobacter species in freshwater wetlands.

The aim of this study was to assess the incidence of Enterobacteriaceae (potential human and animal pathogens) in wetlands. Enterobacteriaceae, selected from the sediments and rhizosphere of wetland plant Juncus effusus L., were analysed using classical microbiological methods, API20E, API20NE, fatty acid analyses, and 16S rRNA sequencing. Assessed virulence factors include antibiotic resistance, presence of plasmids and capsules. Klebsiella pneumoniae, Enterobacter cloacae and Enterobacter asburiae, known human pathogens, were identified. K. pneumoniae 16S rRNA gene sequence showed the significant hit (E < 0.001) with the unculturable bacteria obtained from faeces of elderly individuals (accession number AB099804) when Genbank database was used. Ent. asburiae 16S rRNA gene sequence showed the significant hit with (E < 0.001) with the unculturable bacteria obtained from the pig gastrointestinal tract (accession number AF371852). The rate of antibiotic resistance (<50 microg ml(-1)) was high for ampicillin and cephalosporins for the most strains (75.7%) yet low (>10 to 20 microg ml(-1)) for kanamycin, tetracycline and chloramphenicol for all strains tested. Capsules were detected in all investigated strains. PCR detected membrane protein but not chromosomally encoded beta-lactamase. The antibiotic resistance of tested strains and presence of capsules (protect micro-organisms from phagocytosis) suggest that wetland sediments and rhizosphere present a potential reservoirs for enteric human and animal pathogens.

A simultaneous outbreak on a neonatal unit of two strains of multiply antibiotic resistant Klebsiella pneumoniae controllable only by ward closure

Two aminoglycoside-resistant strains of Klebsiella pneumoniae caused an outbreak on the neonatal unit at St Thomas' Hospital. One, which affected 18 patients, was capsular type K18 and resistant to newer cephalosporins by the production of the extended-spectrum β-lactamase SHV-2; the other, which colonized four patients, was capsular non-typeable and did not produce extended-spectrum β-lactamase. Both strains were probably brought into the unit by carrier patients; the probable carrier of the non-typeable strain was transferred from another hospital but was negative on a single admission screen; the probable carrier of the K18 strain was not screened on admission because he had been born at St Thomas', but his mother had been transferred from another hospital. Despite intensive efforts to control the outbreak by standard methods of hand washing, screening, patient isolation and environmental cleaning, a total of 22 neonates on the unit eventually became colonized or infected. One of three patients with bacteraemia died. A small proportion of samples of expressed breast milk, electronic thermometers and oxygen saturation probes were contaminated by the K18 strain and may have contributed to some of the cross-infection, but this did not explain the extent of the outbreak. The outbreak was controlled only by opening a temporary ward for colonized neonates and another for newly born babies, which allowed the closure and cleaning of the main neonatal unit. Multiply antibiotic resistant klebsiellas may be highly epidemic and cause serious, difficult-to-control outbreaks on neonatal units. All patients, regardless of their admission history, should be screened on admission for carriage of multiply resistant enterobacteria by a sensitive method, and units should have plans for temporary ward closure should outbreaks occur.

Membrane Permeability Modifications Are Involved in Antibiotic Resistance in Klebsiella pneumoniae

Two Klebsiella pneumoniae strains selected according to their high cross-resistance pattern to cephalosporins were characterized by (i) outer membrane protein content such as OmpA or nonspecific porins, (ii) MICs of various cephalosporins and unrelated antibiotics, (iii) β-lactamase production, and (iv) active efflux of fluoroquinolones. An association of porin deficiency and β-lactamase production induced a noticeable cephalosporin resistance. In addition to these mechanisms, the presence of an active efflux participating in high-level fluoroquinolone resistance was identified in one strain. The decrease of antibiotic uptake associated with efflux explains the Klebsiella adaptation against the drugs present in the environment.

Cooperative evolution of mechanisms of β-lactam resistance

Cooperative evolution of mechanisms of β-lactam resistance

Polymorphonuclear leukocyte transfusion for the treatment of sepsis in the newborn infant

A therapeutic trial of transfusions with polymorphonuclear leukocyte concentrates was performed in newborn infants with bacterial sepsis proven by blood culture. With each transfusion, 20 ml/kg of a preparation obtained by continuous flow filtration leukapheresis, and containing 0.5 to 1 x 10(9) WBC, with less than 6% lymphocytes, was administered. Twenty newborn infants with sepsis received from 2 to 15 PMN transfusions. Results were compared with findings in 18 newborn infants with sepsis admitted during the trial period, and not treated because of unavailability of the PMN preparation (Group B). Infants with fulminant illness were excluded from both groups. Groups A and B were similar with respect to clinical characteristics and to etiology (in the majority cases a highly antibiotic-resistant Klebsiella). The mortality rate was significantly lower in Group A than in Group B in the whole series (10% vs 72%, P < 0.001), and also in the subgroups with birth weight equal or below 1,500 gm (10% vs 91%, P < 0.001). Major complications and associated conditions (i.e., necrotizing enterocolitis, meningitis, pneumonia, peritonitis, osteoarthritis, disseminated intravascular coagulation) were observed in 12 patients of Group B, and in only three infants of Group A. Untoward effects attributable to PMN transfusions were never observed. PMN transfusion was a highly effective therapeutic tool in our population of infected newborn infants.

R-factor responsible for an outbreak of multiply antibiotic-resistant Klebsiella pneumoniae

Seven serotypes of Klebsiella pneumoniae isolated from different patients demonstrated resistance to the same eight antibiotics. A plasmid carrying resistance determinants to these antibiotics and mercury salts could be transferred in toto to a plasmidless strain of Escherichia coli. All E. coli transconjugants showed the same antibiotic resistance pattern. Digestion with restriction endonucleases yielded patterns that were identical for each of the R-factor transferred from the multiply resistant serotypes. Moreover, deoxyribonucleic acid-deoxyribonucleic acid hybridization demonstrated identity between the probe, pMAC20 (an R-factor from one serotype), and all R-factors isolated from the multiply resistant strains of K. pneumoniae and the E. coli transconjugants tested.

POLYMORPHONUCLEAR LEUKOCYTES (PMN) TRANSFUSION FOR THE TREATMENT OF NEONATAL SEPSIS

25 newborns with bacterial sepsis received from 2 to 14 transfusions (20 ml/kg of a suspension containing .5-1 × 109 PMNs, and less than 6% lymphocytes, obtained by continuous flow filtration leukapheresis). Results were compared with findings in 40 septic newborns not treated by PMNs transfusions (GROUP B). GROUPS A and B were comparable with respect to clinical characteristics and etiology (in most cases a highly antibiotic-resistant Klebsiella). The mortality rate was lower in GROUP A than in GROUP B (8% vs 70%, p 1500 g (8% vs 50%, p = 0.02). Major complications and associated conditions (i.e. NEC, meningitis, pneumonia, peritonitis, osteoarthritis, DIC) were observed in 18 patients of GROUP B, and only in 3 of GROUP A. Untoward effects attributable to the PMNs transfusion were never observed. It is concluded that PMNs transfusion represents an highly effective treatment of neonatal sepsis. (C.N.R. project MPP3, n. 79.010.022.83).

Analysis of Increasing Antibiotic Resistance of Klebsiella pneumoniae Relative to Changes in Chemotherapy

Three antibiotic-resistant strains of Klebsiella pneumoniae responsible for outbreaks of systemic infection in a hospital nursery were analyzed. Each organism emerged after prolonged use of a drug to which it was resistant. The first strain (RO16) produced neomycin phosphotransferase and contained three plasmids with molecular weights of 24, 25, and 30 X 10(6) daltons, respectively. Resistance to ampicillin, carbenicillin, neomycin, and kanamycin was transferred by conjugation at a frequency of 10(-6). The second strain of K. pneumoniae (RO106) produced gentamicin adenyltransferase and maintained aminoglycoside resistance only when propagated in antibiotic-containing medium. DNA analyses revealed eight species of plasmid DNA: one species with a molecular weight of 70 X 10(6) daltons apparently accounted for conjugal transfer of resistance to ampicillin, carbenicillin, chloramphenicol, and streptomycin. The third strain (RO180) was resistant primarily to colistin and lacked plasmids. Control of the outbreak due to this strain was achieved by aminoglycoside therapy.

Transferable antibiotic resistance (R factors) in a newborn ward.

Antibiotic-resistant klebsiella, enterobacter and pseudomonas strains were isolated from nose and throat swabs of the patients in a newborn ward. Most of the patients received antibiotic treatment, while only few had manifest infection. R factors transferring resistance to several drugs were detected in 2 of the 26 klebsiella-enterobacter but in none of the 10 pseudomonas strains. The R factors occurred among strains with multiple resistance. The demonstration of R factors among these opportunistic bacteria adds to reasons against liberal use of antibiotics.

Transfer of antibiotic resistance (R factor) in the mouse intestine.

SummaryAn antibiotic resistant Klebsiella pneumoniae containing an R factor was fed to germ-free mice previously colonized with an antibiotic susceptible E. coli. Within a month, 21% of the E. coli acquired multiple drug resistance without the selective pressure of antibiotic therapy. However, the rate of spread of the R factor was significantly slower than in vitro.