LysR-type transcriptional regulators are involved in the regulation of numerous cellular metabolic processes in Klebsiella pneumoniae, leading to severe infection. Earlier, we found a novel LysR family gene, named kp05372, in a strain of K. pneumoniae (designated GPKP) isolated from forest musk deer. To study the function of this gene in relation to the biological characteristics of GPKP, we used the suicide plasmid and conjugative transfer methods to construct deletion mutant strain GPKP-Δkp05372; moreover, we also constructed the GPKP-Δkp05372+ complemented strain. The role of this gene was determined by comparing the following characteristics of three strains: growth curves, biofilm formation, drug resistance, stress resistance, median lethal dose (LD₅₀), organ colonization ability, and the histopathology of GPKP. Real-time polymerase chain reaction (RT-PCR) was used to test the expression level of seven genes upstream of kp05372. There was no significant difference in the growth rates when comparing the three bacterial strains, and no significant difference was recorded at different osmotic pressures, temperatures, salt contents, or hydrogen peroxide concentrations. The GPKP-Δkp05372 mutant formed a weak biofilm, and the other two strains formed medium biofilm. The drug resistance of the GPKP-Δkp05372 mutant toward cephalothin, cotrimoxazole, and polymyxin B was changed. The acid tolerance of the deletion strain was stronger than that of the other two strains. The LD₅₀ values of the wild-type and complemented strains were 174-fold and 77-fold higher than that of the GPKP-Δkp05372 mutant, respectively. The colonization ability of the GPKP-Δkp05372 mutant in the heart, liver, spleen, kidney, and intestine was the weakest. The three strains caused different histopathological changes in the liver and lungs. In the GPKP-Δkp05372 mutant, the relative expression levels of kp05374 and kp05379 were increased to 1.32-fold and 1.42-fold, respectively, while the level of kp05378 was decreased by 42%. Overall, the deletion of kp05372 gene leads to changes in the following: drug resistance and acid tolerance; decreases in virulence, biofilm formation, and colonization ability of GPKP; and regulation of the upstream region of adjacent genes.
Animals ; Bacterial Proteins/physiology* ; Biofilms ; Deer/microbiology* ; Drug Resistance, Bacterial ; Female ; Klebsiella Infections/pathology* ; Klebsiella pneumoniae/growth & development* ; Male ; Mice ; Transcription Factors/physiology*

OBJECTIVETo characterize carbapenem (CPM)-non-susceptible Klebsiella pneumoniae (K. pneumoniae) and carbape-nemase produced by these strains isolated from Beijing Children's Hospital based on a five-year surveillance.

METHODSThe Minimal Inhibition Concentration values for 15 antibiotics were assessed using the Phonix100 compact system. PCR amplification and DNA sequencing were used to detect genes encoding carbapenemases. WHONET 5.6 was finally used for resistance analysis.

RESULTSIn total, 179 strains of CPM-non-susceptible K. pneumoniae were isolated from January, 2010 to December, 2014. The rates of non-susceptible to imipenem and meropenem were 95.0% and 95.6%, respectively. In the 179 strains, 95 (53.1%) strains carried the blaIMP gene, and IMP-4 and IMP-8 were detected in 92 (96.8%) and 3 (3.2%) IMP-producing isolates, respectively. 65 (36.3%) strains carried the blaNDM-1 gene. 6 (3.4%) strains carried the blaKPC gene, and KPC-2 were detected in 6 KPC-producing isolates. In addition, New Delhi-Metallo-1 (NDM-1) producing isolates increased from 7.1% to 63.0% in five years and IMP-4 producing isolates decreased from 75.0% to 28.3%.

CONCLUSIONHigh frequencies of multiple resistances to antibiotics were observed in the CPM-non-susceptible K. pneumoniae strains isolated from Beijing Children's Hospital. The production of IMP-4 and NDM-1 metallo-β-lactamases appears to be an important mechanism for CPM-non- susceptible in K. pneumoniae.

Anti-Bacterial Agents ; pharmacology ; Bacterial Proteins ; genetics ; metabolism ; Child ; China ; epidemiology ; Drug Resistance ; Gene Expression Regulation, Bacterial ; physiology ; Gene Expression Regulation, Enzymologic ; physiology ; Hospitals, Pediatric ; Humans ; Klebsiella Infections ; epidemiology ; microbiology ; Klebsiella pneumoniae ; drug effects ; enzymology ; genetics ; Microbial Sensitivity Tests ; Population Surveillance ; Time Factors ; beta-Lactamases ; genetics ; metabolism

Carbapenems ; pharmacology ; Drug Resistance, Bacterial ; Humans ; Klebsiella Infections ; drug therapy ; microbiology ; Klebsiella pneumoniae ; drug effects

INTRODUCTIONIncreasing resistance in Escherichia coli and Klebsiella pneumoniae to firstline antibiotics makes therapeutic options for urinary tract infections (UTIs) challenging. This study investigated the in vitro efficacies of 6 antibiotics against multidrug resistant (MDR) uropathogens.

MATERIALS AND METHODSMinimum inhibitory concentrations to ceftibuten, cefpodoxime, fosfomycin, mecillinam, temocillin, and trimethoprim were determined against 155 MDR-isolates of E. coli and K. pneumoniae. The presence of extended-spectrum beta-lactamases (ESBL) and plasmid-borne AmpC enzymes was determined by phenotypic testing with genotyping performed by multiplex polymerase chain reaction.

RESULTSTemocillin demonstrated highest susceptibility rates for both E. coli (95%) and K. pneumoniae (95%) when breakpoints for uncomplicated UTIs were applied; however, temocillin susceptibility was substantially lower when "systemic infection" breakpoints were used. Fosfomycin demonstrated the best in vitro efficacy of the orally available agents, with 78% and 69% of E. coli and K. pneumoniae isolates susceptible, respectively. The next most effective antibiotics were ceftibuten (45%) and mecillinam (32%). ESBL and ampC genes were present in 47 (30%) and 59 (38%) isolates.

CONCLUSIONThis study demonstrated few oral therapeutic options for MDR-uropathogens, with fosfomycin demonstrating the best in vitro activity.

Amdinocillin ; pharmacology ; Anti-Bacterial Agents ; pharmacology ; Bacterial Proteins ; genetics ; Ceftizoxime ; analogs & derivatives ; pharmacology ; Cephalosporins ; pharmacology ; Drug Resistance, Multiple, Bacterial ; genetics ; Escherichia coli ; drug effects ; genetics ; Escherichia coli Infections ; microbiology ; Fosfomycin ; pharmacology ; Genotype ; Humans ; In Vitro Techniques ; Klebsiella Infections ; microbiology ; Klebsiella pneumoniae ; drug effects ; genetics ; Microbial Sensitivity Tests ; Multiplex Polymerase Chain Reaction ; Penicillins ; pharmacology ; Singapore ; Trimethoprim ; pharmacology ; Urinary Tract Infections ; microbiology ; beta-Lactamases ; genetics

To analyze the clinical characteristics of continuous ambulatory peritoneal dialysis (CAPD) associated peritonitis in the tertiary hospitals and to discuss the preventive and therapeutic strategy.
 Methods: The clinical characteristics, pathogens, resistance and outcomes of 126 CAPD associated peritonitis in 104 patients from Jan, 2013 to June, 2016, were retrospectively analyzed.
 Results: Among the patients, the incidence rates of abdominal pain, fever, diarrhea and emesis were 104 (82.54%), 56 (44.44%), 49 (38.89%), and 31 (23.60%), respectively. Among them, 88 patients suffered peritonitis once, other 16 patients suffered multiple peritonitis or recurrent peritonitis for 38 times. Among the 38 times, the numbers for recurrent, repeated or catheter-associated peritonitis were 2, 2, or 3, respectively. Peritoneal fluids from 103 cases were cultured, and 64 cases were positive in bacteria, with a rate of 62.14%. A total of 70 strains of bacteria were separated, including 42 strains of gram-positive bacteria, 21 strains of gram-negative bacteria, and 7 strains of fungus. The most common gram-positive pathogens were Staphylococcus epidermidis, Enterococcus faecalis and Staphylococcus haemolyticus, while Escherichia coli, Klebsiella pneumoniae and Klebsiella pneumoniae were the most common gram-negative bacteria. Candida albicans was the major fungal pathogens. Gram-positive cocci showed resistance to gentamycin, levofloxacin, moxifloxacin, vancomycin and linezolid, with a rate at 20.00%, 36.11%, 5%, 0%, and 0%, respectively. The gram-negative bacilli were resistent to cefoperazone/sulbactam, gentamycin, cephazolin, and ceftazidime, with a rate at 6.25%, 10.53%, 64.29%, and 15.38%, respectively. There were no imipenem, amikacin, piperacillin/tazobactam-resistant strains were found.
 Conclusion: The most common pathogen causing CAPD associated peritonitis is gram-positive bacteria. It is crucial to take the anti-infection therapy for CAPD associated peritonitis early. The positive rates for bacterial culture need to be enhanced through improvement of methods. At the same time, doctors could improve the outcome of CAPD associated peritonitis by adjusting the medication according to the drug sensitivity results.
Abdominal Pain ; epidemiology ; Anti-Bacterial Agents ; Bacteria ; Bacterial Infections ; epidemiology ; microbiology ; Candidiasis ; epidemiology ; Catheters ; adverse effects ; microbiology ; Diarrhea ; epidemiology ; Drug Resistance, Bacterial ; Enterococcus faecalis ; Escherichia coli ; Fever ; epidemiology ; Gram-Negative Bacteria ; Gram-Positive Bacteria ; Humans ; Imipenem ; Klebsiella pneumoniae ; Microbial Sensitivity Tests ; Mycoses ; epidemiology ; Penicillanic Acid ; analogs & derivatives ; Peritoneal Dialysis ; Peritoneal Dialysis, Continuous Ambulatory ; adverse effects ; Peritonitis ; complications ; epidemiology ; microbiology ; Piperacillin ; Piperacillin, Tazobactam Drug Combination ; Recurrence ; Retrospective Studies ; Staphylococcus epidermidis ; Staphylococcus haemolyticus ; Vomiting ; epidemiology

No abstract available.
Aged ; Female ; Humans ; Klebsiella Infections/*diagnosis/microbiology ; Klebsiella pneumoniae/*enzymology/isolation & purification ; Pneumonia/*diagnosis/microbiology ; Turkey ; beta-Lactamases/*metabolism

BACKGROUND: The emergence of carbapenem-resistant Klebsiella pneumoniae poses a serious problem to antibiotic management. We investigated the beta-lactamases in a group of carbapenem-resistant K. pneumoniae clinical isolates from Turkey. METHODS: Thirty-seven strains of K. pneumoniae isolated from various clinical specimens were analyzed by antimicrobial susceptibility testing, PCR for the detection of beta-lactamase genes, DNA sequencing, and repetitive extragenic palindronic (REP)-PCR analysis. RESULTS: All 37 isolates were resistant to ampicillin, ampicillin/sulbactam, piperacillin, piperacillin/tazobactam, ceftazidime, cefoperazone/sulbactam, cefepime, imipenem, and meropenem. The lowest resistance rates were observed for colistin (2.7%), tigecycline (11%), and amikacin (19%). According to PCR and sequencing results, 98% (36/37) of strains carried at least one carbapenemase gene, with 32 (86%) carrying OXA-48 and 7 (19%) carrying NDM-1. No other carbapenemase genes were identified. All strains carried a CTX-M-2-like beta-lactamase, and some carried SHV- (97%), TEM- (9%), and CTX-M-1-like (62%) beta-lactamases. Sequence analysis of bla(TEM) genes identified a bla(TEM-166) with an amino acid change at position 53 (Arg53Gly) from bla(TEM-1b), the first report of a mutation in this region. REP-PCR analysis revealed that there were seven different clonal groups, and temporo-spatial links were identified within these groups. CONCLUSIONS: Combinations of beta-lactamases were found in all strains, with the most common being OXA-48, SHV, TEM, and CTX-M-type (76% of strains). We have reported, for the first time, a high prevalence of the NDM-1 (19%) carbapenemase in carbapenem-resistant K. pneumoniae from Turkey. These enzymes often co-exist with other beta-lactamases, such as TEM, SHV, and CTX-M beta-lactamases.
Anti-Bacterial Agents/*pharmacology ; Bacterial Proteins/*genetics/metabolism ; Carbapenems/*pharmacology ; DNA, Bacterial/chemistry/genetics/metabolism ; Drug Resistance, Bacterial ; Genotype ; Humans ; Klebsiella Infections/diagnosis/microbiology ; Klebsiella pneumoniae/*drug effects/enzymology/isolation & purification ; Microbial Sensitivity Tests ; Polymerase Chain Reaction ; Sequence Analysis, DNA ; Turkey ; beta-Lactamases/*genetics/metabolism

INTRODUCTIONTreatment of acute lymphoblastic leukaemia (ALL) using intensive chemotherapy has resulted in high cure rates but also substantial morbidity. Infective complications represent a significant proportion of treatment-related toxicity. The objective of this study was to describe the microbiological aetiology and clinical outcome of episodes of chemotherapy-induced febrile neutropaenia in a cohort of children treated for ALL at our institution.

MATERIALS AND METHODSPatients with ALL were treated with either the HKSGALL93 or the Malaysia-Singapore (Ma-Spore) 2003 chemotherapy protocols. The records of 197 patients who completed the intensive phase of treatment, defined as the period of treatment from induction, central nervous system (CNS)-directed therapy to reinduction from June 2000 to January 2010 were retrospectively reviewed.

RESULTSThere were a total of 587 episodes of febrile neutropaenia in 197 patients, translating to an overall rate of 2.98 episodes per patient. A causative pathogen was isolated in 22.7% of episodes. An equal proportion of Gram-positive bacteria (36.4%) and Gram-negative bacteria (36.4%) were most frequently isolated followed by viral pathogens (17.4%), fungal pathogens (8.4%) and other bacteria (1.2%). Fungal organisms accounted for a higher proportion of clinically severe episodes of febrile neutropaenia requiring admission to the high-dependency or intensive care unit (23.1%). The overall mortality rate from all episodes was 1.5%.

CONCLUSIONFebrile neutropaenia continues to be of concern in ALL patients undergoing intensive chemotherapy. The majority of episodes will not have an identifiable causative organism. Gram-positive bacteria and Gram-negative bacteria were the most common causative pathogens identified. With appropriate antimicrobial therapy and supportive management, the overall risk of mortality from febrile neutropaenia is extremely low.

Candidiasis ; epidemiology ; Chemotherapy-Induced Febrile Neutropenia ; epidemiology ; microbiology ; Child ; Cohort Studies ; Escherichia coli Infections ; epidemiology ; Gram-Negative Bacterial Infections ; epidemiology ; Gram-Positive Bacterial Infections ; epidemiology ; Humans ; Influenza, Human ; epidemiology ; Klebsiella Infections ; epidemiology ; Mycoses ; epidemiology ; Precursor Cell Lymphoblastic Leukemia-Lymphoma ; drug therapy ; Pseudomonas Infections ; epidemiology ; Retrospective Studies ; Singapore ; epidemiology ; Staphylococcal Infections ; epidemiology ; Virus Diseases ; epidemiology

The emergence and spread of carbapenems-resistant Klebsiella pneumoniae (CRKP) in burn ward is an important threat to burn management. CRKP isolates are resistant to almost all available antibiotics and are susceptible only to polymyxins and tigecycline. The mechanism of the drug resistance of CRKP is associated with the plasmid-encoded carbapenemase Klebsiella pneumoniae carbapenemase (KPC), a carbapenem-hydrolyzing β-lactamase. Antibiotics which can currently be used to treat CRKP infection include polymyxins, tigecycline, and some aminoglycosides. The efficacy of using antibiotics in combination is better than that of single-agent therapy for the treatment of CRKP infection in bloodstream. In order to control CRKP infection in burn patients, strategies for preventing CRKP dissemination in burn ward are strongly advocated.
Anti-Bacterial Agents ; therapeutic use ; Bacterial Proteins ; Burns ; drug therapy ; Carbapenems ; pharmacology ; Drug Resistance, Bacterial ; Humans ; Klebsiella Infections ; drug therapy ; microbiology ; prevention & control ; Klebsiella pneumoniae ; drug effects ; Microbial Sensitivity Tests ; Minocycline ; analogs & derivatives ; therapeutic use ; beta-Lactam Resistance ; beta-Lactamases

The emergence of multidrug-resistant Klebsiella pneumoniae highlights the need to develop preventive measures to ameliorate Klebsiella infections. Bacteria-derived extracellular vesicles (EVs) are spherical nanometer-sized proteolipids enriched with outer membrane proteins. Gram-negative bacteria-derived EVs have gained interest for use as nonliving complex vaccines. In the present study, we evaluated whether K. pneumoniae-derived EVs confer protection against bacteria-induced lethality. K. pneumoniae-derived EVs isolated from in vitro bacterial culture supernatants induced innate immunity, including the upregulation of co-stimulatory molecule expression and proinflammatory mediator production. EV vaccination via the intraperitoneal route elicited EV-reactive antibodies and interferon-gamma-producing T-cell responses. Three vaccinations with the EVs prevented bacteria-induced lethality. As verified by sera and splenocytes adoptive transfer, the protective effect of EV vaccination was dependent on both humoral and cellular immunity. Taken together, these findings suggest that K. pneumoniae-derived EVs are a novel vaccine candidate against K. pneumoniae infections.
Animals ; Bacterial Vaccines/immunology/*microbiology/*therapeutic use ; Extracellular Vesicles/immunology/*microbiology ; Female ; Humans ; Immunity, Cellular ; Immunity, Innate ; Interferon-gamma/immunology ; Klebsiella Infections/immunology/*prevention &control ; Klebsiella pneumoniae/*immunology ; Mice, Inbred C57BL ; Vaccination