BACKGROUND/AIMS: The number of urinary tract infections (UTIs) caused by extended-spectrum beta-lactamase-producing Escherichia coli (ESBL-EC) is increasing. In an outpatient setting, there are limited therapeutic options to treat ESBL-producing pathogens. We evaluated the outcomes of amikacin outpatient parenteral antibiotic therapy (OPAT) for UTIs caused by ESBL-EC in patients not pre-treated with carbapenem. METHODS: We retrospectively evaluated the outcomes of amikacin OPAT for UTIs caused by ESBL-EC. RESULTS: From November 2011 to October 2012, eight females, who could not be hospitalized for carbapenem treatment, were treated with amikacin OPAT for nine episodes of non-bacteremic ESBL-EC UTIs. Seven of the eight patients had one or more comorbidities. Of the nine UTI cases, three had symptomatic lower UTIs and six had non-bacteremic upper UTIs. In all of the cases, symptomatic and laboratory improvements were observed following amikacin OPAT. One patient showed a delayed relapse with bilateral microabscesses 3 weeks after treatment cessation; however, a clinical and microbiological cure was eventually reached. All of the patients were able to tolerate amikacin OPAT without any significant nephrotoxicity or ototoxicity. CONCLUSIONS: Amikacin OPAT represents a feasible therapeutic option for non-bacteremic UTIs caused by ESBL-EC in settings with limited resources.
Adult ; Aged ; Aged, 80 and over ; Ambulatory Care ; Amikacin/administration & dosage/adverse effects/*therapeutic use ; Drug Administration Schedule ; Escherichia coli/*drug effects/enzymology/isolation & purification ; Escherichia coli Infections/diagnosis/*drug therapy/microbiology/urine ; Humans ; Microbial Sensitivity Tests ; Middle Aged ; Recurrence ; Remission Induction ; Retrospective Studies ; Time Factors ; Treatment Outcome ; Urinalysis ; Urinary Tract Infections/diagnosis/*drug therapy/microbiology/urine ; Urine/microbiology ; beta-Lactamase Inhibitors/administration & dosage/adverse effects/*therapeutic use ; beta-Lactamases/*metabolism

The use of intrauterine devices (IUDs) have been widespread since the 1960s. In 2002, the World Health Organization estimated that approximately 160 million women worldwide use IUDs. However, IUDs are associated with short-term complications such as vaginal bleeding, pelvic discomfort, dyspareunia and pelvic infection. Herein, we report the case of a woman who had recurrent urinary tract infection (UTI) due to the use of an IUD, even after treatment. The patient developed four episodes of UTI within a seven-month period after IUD insertion. During each episode of UTI, extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli (E. coli) was cultured from the patient’s midstream urine. The IUD was finally removed, and culture of the removed IUD was positive for ESBL-producing E. coli. An infected IUD as a source of recurrent UTI should be considered in women with IUD in situ who develop recurrent UTI even after treatment.
Adult ; Anti-Bacterial Agents ; therapeutic use ; Escherichia coli Infections ; drug therapy ; etiology ; Female ; Humans ; Intrauterine Devices ; adverse effects ; Microbial Sensitivity Tests ; Recurrence ; Treatment Outcome ; Urinary Tract Infections ; drug therapy ; etiology ; microbiology ; Uropathogenic Escherichia coli ; enzymology ; beta-Lactamases ; metabolism

The tissue-specific and MeJA-induced transcriptional levels of BcUGT3 and BcUGT6 in Bupleurum chinense were analyzed in the present study. The transcriptional levels of BcUGT3 in root, leaf, flower and fruit were similar and they all were higher than those in stem. The transcriptional level of BcUGT6 was the highest in leaf and the lowest in flower among in all tested tissues. With non-treated adventitious roots as control, BcUGT6's transcriptional levels were elevated to nearly 2 folds for 2 h, 8 h, 24 h, 2 d and 4 d in MeJA-treated adventitious roots of B. chinense. It showed that the transcriptional level of BcUGT6 was slightly affected by MeJA. While, BcUGT3's transcriptional levels were gradually elevated, and till 4 d after MeJA treatment, the expression level was about 7 folds than that of non-treated control. Using pET-28a (+), the expressions of two genes was investigated. Induced by IPTG, the target proteins were expressed in E. coli and then purified. All the results obtained in the present study will be helpful for follow-up bio-function analysis of BcUGT3 and BcUGT6.
Acetates ; pharmacology ; Bupleurum ; cytology ; enzymology ; genetics ; Cell Membrane ; metabolism ; Cyclopentanes ; pharmacology ; Escherichia coli ; genetics ; Gene Expression ; Gene Expression Regulation, Plant ; drug effects ; Hexosyltransferases ; chemistry ; genetics ; isolation & purification ; metabolism ; Intracellular Space ; metabolism ; Oxylipins ; pharmacology ; Protein Sorting Signals ; Protein Structure, Secondary ; Protein Transport ; Sequence Analysis ; Transcription, Genetic ; drug effects

OBJECTIVETo construct plant expression pCAMBIA1301-PMI by substituting PMI for hygromycin resistance gene in pCAMBIA1301 and obtain transgenic Salvia miltiorrhiza f. alba using PMI-mannose selection system.

METHODThe 6-phosphomannose isomerase gene (PMI) of Escherichia coli was amplified by PCR. Sequence analysis showed that it shared 100% amino acids identities with the sequences of PMI genes isolates reported in the NCBI. Based on pCAMBIA1305, the plant expression pCAMBIA1305-PMI was constructed successfully by substituting PMI for hygromycin resistance gene in pCAMBIA1305. pCAMBIA1305-PMI was transformed into Agrobacterium tumefaciens LBA4404, and then the leaves of S. miltiorrhiza f. alba were inoculated in LBA4404 with pCAMBIA1305-PMI.

RESULTPlant expression pCAMBIA1301-PMI was successfully constructed and the leaves of S. miltiorrhiza f. alba inoculated in LBA4404 with pCAMBIA1305-PMI were selected on medium supplemented with a combination of 20 g x L(-1) mannose and 10 g x L(-1) sucrose as a carbon source. The transformation efficiency rate was 23.7%.

CONCLUSIONGenetic transformation was confirmed by PCR, indicating that a new method for obtaining transgenic S. miltiorrhiza f. alba plants was developed using PMI-mannose selection system.

Anti-Bacterial Agents ; pharmacology ; Biomarkers ; Cinnamates ; pharmacology ; Escherichia coli ; enzymology ; genetics ; Escherichia coli Proteins ; genetics ; metabolism ; Gene Expression ; Genetic Vectors ; genetics ; metabolism ; Hygromycin B ; analogs & derivatives ; pharmacology ; Mannose-6-Phosphate Isomerase ; genetics ; metabolism ; Plants, Genetically Modified ; drug effects ; genetics ; metabolism ; Salvia miltiorrhiza ; drug effects ; genetics ; metabolism ; Transformation, Genetic

OBJECTIVETo evaluate the antimicrobial activity of total alkaloids extracted from Sophorea alopecuroides L. (TASA) against clinical isolated extended-spectrum beta-lactamases (ESBLs) producing Escherichia coli (E. coli) strains.

METHODSThe antibacterial activity of TASA either itself or in combination with cefotaxime (CTX) or ceftazidime (CAZ) was investigated by using the microbroth dilution method and phenotypic confirmatory disk diffusion test against three clinical isolated ESBLs-producing E. coli strains; the interactions of TASA and CTX or CAZ were ascertained by evaluating the fractional inhibitory concentration index (FICI).

RESULTSThe antibacterial activity of either TASA itself or in combination with CTX or CAZ was found. The minimum inhibitory concentration (MICs) of TASA against the ESBLs producing isolates was 12.5 mg/mL. In the combinations with a sub-inhibitory concentration of TASA, a synergistic effect on CTX and CAZ against the ESBLs producing isolates was observed. Similarly, the isolates exposed to lower dose of TASA yielded an increased susceptibility to CTX and CAZ by 8-16 folds determined by microdilution assay. Moreover, enzymatic detection of ESBLs demonstrated that TASA induced reversal resistance to CTX and CAZ partially by a mechanism of inhibition of ESBLs activity in these isolates. Additionally, in the tested isolates following the exposure of TASA, molecular analysis verified the SHV-type beta-lactamase encoding ESBL gene in these isolates, and no mutation was introduced into the ESBL gene.

CONCLUSIONSThese results suggest that TASA could be used as a source of natural compound with pharmacological activity of reversal resistance to antimicrobial agent. These findings also indicated that the application of the TASA in combination with antibiotics might prove useful in the control and treatment of infectious diseases caused by the ESBLs producing enterobacteriaceae.

Alkaloids ; isolation & purification ; pharmacology ; Anti-Bacterial Agents ; pharmacology ; Base Sequence ; Cefotaxime ; pharmacology ; Ceftazidime ; pharmacology ; DNA Primers ; Escherichia coli ; drug effects ; enzymology ; Microbial Sensitivity Tests ; Polymerase Chain Reaction ; Sophora ; chemistry ; beta-Lactamases ; biosynthesis

OBJECTIVETo explore the effects of exogenous carbon monoxide-releasing molecules 2 (CORM-2) on the vitality and toxicity of E. coli ATCC 25922, and to analyze the potential mechanism.

METHODS(1) In vitro experiments. Standard strains of E. coli ATCC 25922 were divided into groups A (without addition), B, C, D, and E according to the random number table, and then the latter 4 groups were respectively cultured with 1.2 mmol/L CORM-2, 1.6 mmol/L CORM-2, 1.2 mmol/L inactive CORM-2 (iCORM-2), 1.6 mmol/L iCORM-2, with six samples in each group. After being cultured for 0, 3, 5, 8, 10, 12, 16, 20, 24, 27, 30, 48 hours, proliferative vitality of E. coli was examined (denoted as absorption value under 600 nm wavelength), and bacteria number was counted. Other standard strains of E. coli ATCC 25922 were divided into groups F (without addition) and G (cultured with 0.8 mmol/L CORM-2), the expressions of genes fliA, dnaK, marA, and waaQ related to E. coli were detected by quantitative real-time (qRT)-PCR. (2) In vivo experiments. Other standard strains of E. coli ATCC 25922 were grouped as A', B', C', D', and E' and treated with the same method as that in groups A, B, C, D, and E, and 0.5 mL bacterial liquid of each group were collected when the absorption value of bacterial liquid in group A' was equal to 0.4 (under 600 nm wavelength). Seventy-two C57BL/6 mice were divided into groups, namely blank control (without treatment), H, I, J, K, and L according to the random number table, with 12 mice in each group. The mice in the latter 5 groups were intraperitoneally injected with 0.5 mL bacterial suspension of groups A', B', C', D', and E' respectively. After injection, general condition of mice in groups H, I, J, K, and L was observed. The serum levels of TNF-α and IL-6 were determined at post injection hour (PIH) 6, 12. The liver and lung samples were harvested for determination of myeloperoxidase (MPO) activity at PIH 12. The same process was carried out in blank control group. Data were processed with repeated measure analysis of variance (ANOVA), factorial design ANOVA, one-way ANOVA, and t test.

RESULTS(1) In vitro experiments. Compared with those of groups A and D, the proliferative vitality and bacteria number of E. coli in group B were all decreased (with F values respectively 1170.80, 217.52, P values all below 0.01). Compared with those of groups A and E, the proliferative vitality and bacteria number of E. coli in group C were also obviously decreased (with F values respectively 7948.34, 14 432.85, P values all below 0.01). Compared with those in group F, the expression of fliA was downregulated, while the expressions of dnaK, marA, and waaQ were upregulated in group G (with t values 30.28, -165.54, -168.88, -187.28, P values all below 0.01). (2) In vivo experiments. Symptoms including listlessness and tachypnea were observed in mice in groups H, K, and L, and they were ameliorated or not obvious in groups I and J. At PIH 6, the serum levels of TNF-α and IL-6 in groups H and K were respectively (647.3 ± 3.8) pg/mL, (3.44 ± 0.22) ng/mL and (639.3 ± 0.8) pg/mL, (2.47 ± 0.32) ng/mL, which were obviously higher than those in group I [(124.6 ± 10.7) pg/mL, (1.03 ± 0.16) ng/mL, with t values from 15.22 to 84.03, P values all below 0.01]. The serum levels of TNF-α and IL-6 in group J at PIH 6, 12 were also obviously decreased as compared with those in groups H and L (with t values from 19.27 to 245.34, P values all below 0.01). MPO activity of liver and lung tissues were significantly attenuated in group I at PIH 12 as compared with those in groups H and K, and it was also attenuated in group J when compared with those in groups H and L (with t values respectively from 17.36 to 18.92 and 2.35 to 3.61, P values all below 0.01).

CONCLUSIONSCORM-2 can obviously inhibit the vitality and toxicity of E. coli, which might be attributable to regulation of expressions of genes fliA, dnaK, marA, and waaQ of E. coli.

Animals ; Carbon Monoxide ; metabolism ; DNA-Binding Proteins ; metabolism ; Escherichia coli ; drug effects ; metabolism ; physiology ; Escherichia coli Proteins ; metabolism ; Glycosyltransferases ; metabolism ; HSP70 Heat-Shock Proteins ; metabolism ; Interleukin-6 ; blood ; Liver ; enzymology ; Lung ; enzymology ; Mice ; Mice, Inbred C57BL ; Organometallic Compounds ; pharmacology ; Peroxidase ; metabolism ; Sigma Factor ; metabolism ; Tumor Necrosis Factor-alpha ; blood

We report a case of CTX-M-55-type extended-spectrum beta-lactamase (ESBL)-producing Shigella sonnei infection in a 27-year-old Korean woman who had traveled to China. The patient was admitted to the hospital due to abdominal pain, watery diarrhea, and fever (39.3degrees C). S. sonnei was isolated from her stool specimens, and the pathogen was found to be resistant to cefotaxime due to CTX-M-55-type ESBL. Insertion sequence (IS)Ecp1 was found upstream of the blaCTX-M-55 gene. The blaCTX-M-55 gene was transferred from the S. sonnei isolate to an Escherichia coli J53 recipient by conjugation. Pulsed-field gel electrophoresis and Southern blotting revealed that the blaCTX-M-55 gene was located on a plasmid of approximately 130 kb.
Adult ; Anti-Bacterial Agents/pharmacology ; Asian Continental Ancestry Group ; Cefotaxime/pharmacology ; China ; Drug Resistance, Bacterial/drug effects ; Dysentery, Bacillary/diagnosis/*microbiology ; Electrophoresis, Gel, Pulsed-Field ; Escherichia coli/metabolism ; Feces/microbiology ; Female ; Humans ; Plasmids/chemistry/genetics ; Republic of Korea ; Shigella sonnei/enzymology/*isolation & purification ; Travel ; beta-Lactamases/genetics/*metabolism

We investigated the occurrence and genetic basis of AmpC beta-lactamase (AmpC)-mediated antibiotic resistance, by examining Escherichia coli, Klebsiella pneumoniae, and Proteus mirabilis isolates at a university hospital, from 2007 to 2010. The ampC genes were detected by multiplex AmpC PCR, and AmpC-positive strains were subjected to DNA sequencing. Extended-spectrum beta-lactamase (ESBL) production was assessed using the ESBL disk test based on the utilization of boronic acid. Carbapenem-resistant isolates were further investigated by the modified Hodge test, a carbapenemase inhibition test and SDS-PAGE experiments. AmpC expression was detected in 1.6% of E. coli (39 DHA-1, 45 CMY-2, and 1 CMY-1) isolates, 7.2% of K. pneumoniae (39 DHA-1, 45 CMY-2, and 1 CMY-1) isolates, and 2.5% of P. mirabilis (8 CMY-2 and 1 CMY-1) isolates. Of the 198 acquired AmpC producers, 58 isolates (29.3%) also produced an ESBL enzyme. Among the acquired AmpC-producing K. pneumoniae isolates, the minimum inhibitory concentration (MIC) MIC50/MIC90 values for cefoxitin, cefotaxime, cefepime, imipenem, and meropenem were >32/>32, 16/>32, 1/16, 0.25/0.5, and <0.125/0.125 microg/mL, respectively. The MIC values for carbapenem were > or =2 microg/mL for 2 K. pneumoniae isolates, both of which carried the blaDHA-1 gene with a loss of OmpK36 expression, but were negative for carbapenemase production. The acquisition of AmpC-mediated resistance in K. pneumoniae isolates increased, as did the proportion of AmpC and ESBL co-producers among the hospital isolates. The accurate identification of isolates producing AmpCs and ESBLs may aid in infection control and will assist physicians in selecting an appropriate antibiotic regimen.
Anti-Bacterial Agents/pharmacology ; Bacterial Proteins/*genetics ; DNA, Bacterial/genetics ; Enterobacteriaceae Infections/*epidemiology/*microbiology ; Escherichia coli/drug effects/enzymology/isolation & purification ; Hospitals, University/statistics & numerical data ; Humans ; Klebsiella pneumoniae/drug effects/enzymology/isolation & purification/*physiology ; Microbial Sensitivity Tests ; Multiplex Polymerase Chain Reaction ; Proteus mirabilis/drug effects/enzymology/isolation & purification ; Republic of Korea/epidemiology ; beta-Lactamases/*genetics

OBJECTIVETo study the prevalence and drug resistance of extended-spectrum-β-lactamases (ESBLs)-producing bacteria in blood culture isolated from children with hematological malignancy after chemotherapy.

METHODSBlood samples taken from 3264 children with hematological malignancy and severe infection following chemotherapy between 2002 and 2008 were cultured using the Bact/ALTER 3D blood culture system. VITEK 60 automicroscan was used to identify viral species and to conduct drug resistance tests. The results were indentified according to National Committee for Clinical Laboratory Standard guidelines.

RESULTSFifty-eight strains of Escherichia coli and fifty-one strains of Klebsiella pneumoniae were isolated. Thirty-eight strains of Escherichia coli and nineteen strains of Klebsiella pneumoniae were ESBLs-producing and these ESBLs-producing strains were less susceptible than those that were non-ESBLs-producing to most antibiotics. Both ESBL- and non-ESBL-producing strains were susceptible to imipenem, piperacillin/tazobactam and amikacin.

CONCLUSIONSThe prevalence of ESBLs-producing bacteria is high in childrn with hematological malignancy and infection following chemotherapy. ESBLs-producing bacteria are resistant to common antibiotics, suggesting that antibiotic treatment based on the result of antimicrobial susceptibility test is necessary in these children.

Adolescent ; Bacteremia ; microbiology ; Child ; Child, Preschool ; Drug Resistance, Bacterial ; Escherichia coli ; drug effects ; enzymology ; isolation & purification ; Female ; Hematologic Neoplasms ; drug therapy ; microbiology ; Humans ; Klebsiella pneumoniae ; drug effects ; enzymology ; isolation & purification ; Male ; Microbial Sensitivity Tests ; beta-Lactamases ; biosynthesis

As a large family of hydrolases, GTPases are widespread in cells and play the very important biological function of hydrolyzing GTP into GDP and inorganic phosphate through binding with it. GTPases are involved in cell cycle regulation, protein synthesis, and protein transportation. Chaperones can facilitate the folding or refolding of nascent peptides and denatured proteins to their native states. However, chaperones do not occur in the native structures in which they can perform their normal biological functions. In the current study, the chaperone activity of the conserved GTPases of Escherichia coli is tested by the chemical denaturation and chaperone-assisted renaturation of citrate synthase and α-glucosidase. The effects of ribosomes and nucleotides on the chaperone activity are also examined. Our data indicate that these conserved GTPases have chaperone properties, and may be ancestral protein folding factors that have appeared before dedicated chaperones.
Citrate (si)-Synthase ; chemistry ; Cloning, Molecular ; Conserved Sequence ; Escherichia coli ; cytology ; enzymology ; GTP Phosphohydrolases ; chemistry ; genetics ; isolation & purification ; metabolism ; Guanosine Diphosphate ; pharmacology ; Guanosine Triphosphate ; analogs & derivatives ; pharmacology ; Molecular Chaperones ; chemistry ; genetics ; isolation & purification ; metabolism ; Protein Denaturation ; drug effects ; Protein Renaturation ; drug effects ; Ribosomes ; metabolism ; alpha-Glucosidases ; chemistry