Molecular epidemiology of clinical isolation of carbapenem-resistant Enterobacterales and application of carbapenemase inhibitor enhancement test. / ä¸´åºŠåˆ†ç¦»ç¢³é’éœ‰çƒ¯ç±»è€è¯è‚ æ†èŒç›®ç»†èŒèŒæ ªçš„åˆ†å­æµè¡Œç— å­¦åŠç¢³é’éœ‰çƒ¯é ¶æŠ‘åˆ¶å‰‚å¢žå¼ºè¯•éªŒçš„åº”ç”¨.

OBJECTIVES: The prevalence of carbapenem-resistant Enterobacterales (CRE) presents a significant challenge in clinical anti-infective treatment. This study aims to investigate drug resistance and the molecular epidemiological characteristics of CRE in our area. Additionally, we seek to evaluate practicality of utilizing carbapenemase inhibitor enhancement test in clinical laboratory. METHODS: Non-repeated CREs isolated from clinical specimens at Xiangya Hospital, Central South University, were collected. Minimum inhibitory concentration (MIC) combined with Kirby-Bauer (KB) assay was used to detect the drug susceptibility of the strains, and 13 carbapenemase-producing genes were detected by PCR. The phenotype of 126 strains of carbapenemase-producing Enterobacterales identified by PCR was detected by the carbapenemase inhibitor enhancement test to understand the agreement between the method and the gold standard PCR results. RESULTS: Among 704 CRE strains examined, we observed significant drug resistance in 501 strains dentified as carbapenemase-producing Enterobacterales (CPE). Klebsiella pneumoniae was the predominant CPE strain, followed by Enterobacter cloacae and Escherichia coli. A total of 9 carbapenemase types were detected, including Klebsiella pneumoniae carbapenemase (KPC), New Delhi metallo-ß-lactamase (NDM), Verona integron- encoded metallo-ß-lactamases (VIM), imipenemase (IMP), oxacillinase-48 (OXA-48), and rare imipenem-hydrolyzing ß-lactamase (IMI), adelaide imipenemase (AIM), Bicêtre carbapenemase (BIC), and guiana extended-spectrum ß-lactamase (GES). The detection rate of KPC serine carbapenemase was 61.7% (309/501). The carbapenemase inhibitor enhancement test exhibited a 100% consistency rate for the strains producing Class A serine carbapenemase and/or Class B metallo-ß-lactamases. CONCLUSIONS: CRE strains in Changsha, Hunan, China, are wide distribution and exhibit carbapenemase production. The main mechanism of carbapenem resistance in these bacterias is predominatly attributed to the production of KPC serine carbapenemase. The presence of GES and IMI genes carried by Enterobacterales has been detected for the first time in this region. The carbapenemase inhibitor enhancement test has been proven to be an accurate method for detecting CRE producing Class A serine carbapenemase and/or Class B metallo-ß-lactamases. This method offers simpicity of operation and ease of results interpretation, making it weel-suited meeting the clinical microbiology laboratory's reguirements for the detection of serine carbapenemase and metallo-ß-lactamases.

Clinical value of lymphocyte count in autoimmune diabetic nephropathy. / æ·‹å·´ç»†èƒžè®¡æ•°åœ¨è‡ªèº«å ç–«æ€§ç³–å°¿ç— è‚¾ç— ä¸­çš„ä¸´åºŠä»·å€¼ï¼ˆè‹±æ–‡ï¼‰.

OBJECTIVES: In recent years, the prevalence of diabetic nephropathy (DN) has increased significantly. An increasing number of studies have shown that lymphocyte-associated inflammatory responses play a role in DN. This study aims to investigate the relationship between lymphocytes and DN in patients with autoimmune diabetes. METHODS: The clinical data of 226 patients with Type 1 diabetes (T1D) and 79 patients with latent autoimmune diabetes in adults (LADA) were retrospectively studied and stratified according to the urinary albumin to creatinine ratio (ACR). Risk factors associated with DN were analyzed using correlation analysis and logistic regression. RESULTS: In T1D and LADA patients, systolic blood pressure (SBP), uric acid duration, and diabetes duration in patients with normoalbuminuria were lower or shorter than those in patients with macroalbuminuria (P<0.05). The lymphocyte count of T1D patients was significantly higher than that in LADA patients (P<0.05), while the neutrophil to lymphocyte ratio (NLR) of T1D patients was significantly lower than that in LADA patients (P<0.05). The lymphocyte count in the T1D patients with normoalbuminuria was lower than that those with macroalbuminuria (P<0.05). The NLR was lower in the T1D patients with macroalbuminuria than those with microalbuminuria and normoproteinuria (all P<0.01). Based on logistic regression analysis, lymphocytes were independently associated with DN in T1D after adjusting for various known risk factors such as course of disease, age, gender, dyslipidemia, hypertension, and smoking status. Analysis of the receiver operating characteristic curve of subjects predicting lymphocytes in normoalbuminuria showed that the area under the curve was 0.601 (95% CI 0.510 to 0.693, P=0.039), and when the cutoff value of lymphocytes was 2.332, the sensitivity was 37.0%, and the specificity was 82.5%. CONCLUSIONS: Lymphocyte counts in autoimmune diabetic patients are closely associated with DN, suggesting that lymphocyte-mediated inflammation may be involved in the pathogenesis of DN in autoimmune diabetic patients. This study provides a possible perspective for using lymphocytes as a potential biomarker for the early identification of individuals at risk for DN and potential therapeutic targets for DN.

Molecular epidemiology and clinical characteristics of Clostridioides difficile infection in patients with inflammatory bowel disease from a teaching hospital.

BACKGROUND: Clostridioides difficile infection (CDI) in patients with inflammatory bowel disease (IBD) is of increasing concern. This study aimed to investigate the molecular epidemiology and antimicrobial susceptibilities of toxigenic C. difficile isolated from IBD patients and to evaluate the risk factors for CDI in IBD population. METHODS: Loose or watery stools from IBD patients were tested for glutamate dehydrogenase, C. difficile toxins A&B and anaerobic culture. Toxigenic C. difficile isolates were characterized by multi-locus sequence typing, ribotyping and antimicrobial susceptibility testing. RESULTS: The prevalence of CDI in IBD patients was 13.6% (43/317). The dominant sequence types (STs) were ST35 (20.9%), ST2 (18.6%) and ST37 (16.3%). The most common ribotypes (RTs) were RT 017 (18.6%), RT 012 (14.0%), and RT 220 (14.0%), whereas RT 027 and RT 078 were not detected in this study. All the isolates were susceptible to vancomycin and metronidazole. The multidrug resistance rate of C. difficile RT 017 was higher (p < 0.01) than that of other RT strains. Recent hospitalization, use of corticosteroids and proton pump inhibitors were related to increased risk of CDI in IBD patients; of these, recent hospitalization and proton pump inhibitors use were independent risk factors. CONCLUSION: Patients with IBD have a relatively high incidence rate of CDI. C. difficile RT 017 is most frequently isolated from IBD patients in this region and warrants more attention to its high resistance rate. Clinicians should pay greater attention to CDI testing in IBD patients with diarrhea to ensure early diagnosis and initiation of effective treatment.

Prevalence of Escherichia coli ST1193 Causing Intracranial Infection in Changsha, China.

ST1193 is an emerging new virulent and resistant clone among Escherichia coli with a tendency to spread rapidly across the globe. However, the prevalence of intracranial infection-causing E. coli ST1193 is rarely reported. This study aimed at determining the prevalence of E. coli ST1193 isolates, causing intracranial infections in Changsha, central China. A total of 28 E. coli isolates were collected from the cerebrospinal fluid of patients with intracranial infection over a four-year period. All isolates were differentiated using multilocus sequence typing (MLST), and phylogenetic grouping, and tested for antibiotic resistance. MLST analysis showed 11 sequence types (ST) among the 28 E. coli isolates. The most prevalent ST was B2-ST1193 (28.6%, 8/28), followed by B2-ST131 (21.4%, 6/28) and F-ST648 (10.7%, 3/28). Of the eight ST1193 isolates, three carried CTX-M-55, and one carried CTX-M-27. All eight ST1193 isolates were resistant to Ciprofloxacin, showing gyrA1AB/parC4A mutations. Two ST1193 isolates carried the aac(6')-Ib-cr gene. All ST1193 isolates were recovered from infants with meningitis, with a fatal outcome for one three-month-old infant. ST1193 has emerged as the predominant type of E. coli strain causing intracranial infections in Changsha, China. This study highlights the importance of implementing appropriate surveillance measures to prevent the spread of this emerging public health threat.

Community Fecal Carriage and Molecular Epidemiology of Extended-Spectrum ß-Lactamase- and Carbapenemase-Producing Escherichia coli from Healthy Children in the Central South China.

Background: Fecal carriage of extended-spectrum ß-lactamase-producing Escherichia coli (ESBL-EC) and carbapenemase-producing E. coli (CP-EC) is well reported among hospitalized adults and children. However, there are few studies on the carriage prevalence and ESBL-EC and CP-EC genotypes among healthy children in China. Patients and Methods: Stool samples were collected from 330 students in 2021 from three randomly selected primary schools in Changsha, China. ESBL-EC and CP-EC were screened using CHROMagarTM chromogenic plates. ESBL and carbapenemase production was confirmed using the double-disc synergy test and a modified carbapenem inactivation method, respectively. Antimicrobial susceptibility was tested using the broth microdilution method. Resistance determinants, virulence factors, and phylogenetic groups were determined by PCR and sequencing. Multi-locus sequence typing (MLST) was performed (seven housekeeping genes were amplified and sequenced) on the phylogenic group B2 E. coli to detect high-risk clonal strains such as ST131 E. coli. Then, ST131 E. coli were characterized based on ST131 clades, O-type, and fimH alleles. Results: In total, 118 (35.8%) ESBL-EC and 3 (0.9%) CP-EC were isolated. bla CTX-M was the most common genotype (27.1%), identified in all ESBL-EC, except one, which carried bla SHV-12. One isolate with mcr-1 was found amongst ESBL-EC, whereas all three CP-EC carried bla NDM-1. The predominant sequence type (ST) clones in group B2 were ST131 and ST1193. The prevalence of ST131 E. coli was 9.9%, displaying serotypes O16 and O25b, fimH alleles 30, 41, and 89, and ST131 clades A and C1-M27. Conclusion: In this study, high carriage rate of ESBL-EC was found among healthy children, and the dominant ESBL was CTX-M-14. In addition, high-risk clones (ST131 and ST1193) were also detected. This emphasizes the importance of monitoring ESBL-EC in community settings.

Fructose-coated Ångstrom silver prevents sepsis by killing bacteria and attenuating bacterial toxin-induced injuries.

Serious infection caused by multi-drug-resistant bacteria is a major threat to human health. Bacteria can invade the host tissue and produce various toxins to damage or kill host cells, which may induce life-threatening sepsis. Here, we aimed to explore whether fructose-coated Ångstrom-scale silver particles (F-AgÅPs), which were prepared by our self-developed evaporation-condensation system and optimized coating approach, could kill bacteria and sequester bacterial toxins to attenuate fatal bacterial infections. Methods: A series of in vitro assays were conducted to test the anti-bacterial efficacy of F-AgÅPs, and to investigate whether F-AgÅPs could protect against multi-drug resistant Staphylococcus aureus (S. aureus)- and Escherichia coli (E. coli)-induced cell death, and suppress their toxins (S. aureus hemolysin and E. coli lipopolysaccharide)-induced cell injury or inflammation. The mouse models of cecal ligation and puncture (CLP)- or E. coli bloodstream infection-induced lethal sepsis were established to assess whether the intravenous administration of F-AgÅPs could decrease bacterial burden, inhibit inflammation, and improve the survival rates of mice. The levels of silver in urine and feces of mice were examined to evaluate the excretion of F-AgÅPs. Results: F-AgÅPs efficiently killed various bacteria that can cause lethal infections and also competed with host cells to bind with S. aureus α-hemolysin, thus blocking its cytotoxic activity. F-AgÅPs inhibited E. coli lipopolysaccharide-induced endothelial injury and macrophage inflammation, but not by directly binding to lipopolysaccharide. F-AgÅPs potently reduced bacterial burden, reversed dysregulated inflammation, and enhanced survival in mice with CLP- or E. coli bloodstream infection-induced sepsis, either alone or combined with antibiotic therapy. After three times injections within 48 h, 79.18% of F-AgÅPs were excreted via feces at the end of the 14-day observation period. Conclusion: This study suggests the prospect of F-AgÅPs as a promising intravenous agent for treating severe bacterial infections.

Comparative Evaluation of Seven Tigecycline Susceptibility Testing Methods for Carbapenem-Resistant Enterobacteriaceae.

PURPOSE: Carbapenem-resistant Enterobacteriaceae (CRE) strains are extensively resistant to most antibiotics. Tigecycline is one of the few effective drugs that can be used to treat infections caused by CRE. The aim of this study was to evaluate the accuracy of different methods for detecting the susceptibility of CRE to tigecycline. METHODS: Seven commonly used drug susceptibility testing methods were compared and evaluated for the ability to determine CRE tigecycline susceptibility: broth microdilution (BMD), agar dilution method (ADM), disk diffusion method, Etest, MicroScan, Vitek2 COMPACT, and BD Phoenix 100. RESULTS: The minimum inhibitory concentration (MIC) of tigecycline to inhibit 50% and 90% of CRE growth (MIC50 and MIC90, respectively) assessed by ADM and BD Phoenix 100 was the same as that determined by the reference method, BMD. The MIC50 was 2 µg/mL, and the MIC90 was 4 µg/mL. The highest number of susceptible strains was detected by MicroScan, followed by BMD, Etest, ADM, BD Phoenix 100, Vitek2 COMPACT, and disk diffusion method, in descending order. No significant differences were observed among the tigecycline susceptibility results (P > 0.05) obtained from MicroScan, Etest, BD Phoenix 100, and BMD. BMD confirmed that 82.0% of strains were susceptible to tigecycline. ADM, MicroScan, and BD Phoenix 100 yielded the categorical agreement of 96%, 92%, and 93%, respectively. No method was found to present any very major errors (VMEs), and only the Vitek2 COMPACT yielded major errors (MEs) greater than 3%. CONCLUSION: Among the seven methods tested, the ADM, MicroScan, and BD Phoenix 100 methods were accurate for determining the tigecycline susceptibility of CRE. MicroScan was acceptable with better performance than other methods.

Ångstrom-scale silver particle-embedded carbomer gel promotes wound healing by inhibiting bacterial colonization and inflammation.

Poor wound healing after diabetes or extensive burn remains a challenging problem. Recently, we presented a physical approach to fabricate ultrasmall silver particles from Ångstrom scale to nanoscale and determined the antitumor efficacy of Ångstrom-scale silver particles (AgÅPs) in the smallest size range. Here we used the medium-sized AgÅPs (65.9 ± 31.6 Å) to prepare carbomer gel incorporated with these larger AgÅPs (L-AgÅPs-gel) and demonstrated the potent broad-spectrum antibacterial activity of L-AgÅPs-gel without obvious toxicity on wound healing-related cells. Induction of reactive oxygen species contributed to L-AgÅPs-gel-induced bacterial death. Topical application of L-AgÅPs-gel to mouse skin triggered much stronger effects than the commercial silver nanoparticles (AgNPs)-gel to prevent bacterial colonization, reduce inflammation, and accelerate diabetic and burn wound healing. L-AgÅPs were distributed locally in skin without inducing systemic toxicities. This study suggests that L-AgÅPs-gel represents an effective and safe antibacterial and anti-inflammatory material for wound therapy.

Risk Factors for Subsequential Carbapenem-Resistant Klebsiella pneumoniae Clinical Infection Among Rectal Carriers with Carbapenem-Resistant Klebsiella pneumoniae.

PURPOSE: Carbapenem-resistant Klebsiella pneumoniae (CRKP) infection has become a critical clinical concern for its high mortality. Rectal carriage of CRKP has been reported playing an important role in CRKP infection; however, the extent to which carrier develops clinical CRKP infection is unclear. This study aimed to identify risk factors for developing subsequential CRKP clinical infection in rectal carriers with CRKP. PATIENTS AND METHODS: Patients were screened for rectal carriage of CRKP in a tertiary university hospital; then, rectal CRKP carriers were divided into case group (those who developed subsequential clinical infection) and control group. Demographics, comorbid conditions, invasive procedures, antimicrobial exposure and other clinical parameters of those two groups were compared and analyzed using univariate and multivariate logistic regression analyses. Antimicrobial susceptibility profile and carbapenemase phenotype/genotype of those CRKP isolates were determined. MLST was applied to elucidate the molecular epidemiology of rectal CRKP isolates and clinical infection ones. RESULTS: Eight hundred and thirty-five patients were screened for rectal CRKP carriage. A total of 62 CRKP rectal carriers were identified; among them, 37.1% (23/62) developed CRKP clinical infection. CRKP isolates were resistant to most of the tested antimicrobial agents. ST11 was the dominant MLST type in rectal CRKP isolates (71.0%), and all the 23 clinical infection isolates were ST11. Multivariate analysis revealed that admission to the intensive care unit (ICU) (OR, 6.753; P=0.006), being in coma condition (OR, 11.085; P=0.015) and receiving central venous catheter (OR, 8.628; P=0.003) were independent risk factors for progressing to subsequential CRKP infection among those rectal carriers. CONCLUSION: This study identified independent risk factors for developing subsequential CRKP clinical infection among CRKP rectal carriers, with being in coma condition as a new finding. It would help clinician target those high-risk rectal CRKP-colonized patients for prevention of subsequential clinical infection.

A retrospective study on Escherichia coli bacteremia in immunocompromised patients: Microbiological features, clinical characteristics, and risk factors for shock and death.

BACKGROUND: To evaluate clinical features, bacterial characteristics, and risk factors for shock and mortality of immunocompromised patients with Escherichia coli bacteremia. METHODS: A nearly 6-year retrospective study of E coli bacteremia in 188 immunocompromised patients at Xiangya Hospital was conducted. Demographic, clinical, and laboratory data were documented. Phylogenetic background and virulence factors of E coli isolates were detected by polymerase chain reaction. Risk factors for shock and mortality were also investigated. RESULTS: Of all 188 E coli isolates, most prevalent virulence factors were fimH (91.0%), followed by traT (68.6%) and iutA (67.0%), while papG allele I, gafD, and cdtB were not detected. Phylogenetic group D was dominant (42.0%) among all isolates, and group B2 accounted for 17.6%, while group A and B1 accounted for 28.2% and 12.2%, respectively. In univariate analysis, ibeA and cnf1 were associated with mortality, which were not found in multivariate regression analysis. 22.3% of patients suffered shock, and 30-day mortality rate was 21.3%. MDR (HR 2.956; 95% CI, 1.091-8.012) was the only risk factor for shock, while adult (HR 0.239; 95% CI, 0.108-0.527) was a protective factor. Multivariate analysis revealed that shock (HR 4.268; 95% CI, 2.208-8.248; P < .001) and Charlson index > 2 (HR 2.073; 95% CI, 1.087-3.952; P = .027) were associated with fatal outcome. CONCLUSIONS: Escherichia coli bacteremia was highly lethal in immunocompromised patients, and host-related factors played major roles in poor prognosis, while bacterial determinants had little effect on outcome. This study also provided additional information about the virulence and phylogenetic group characteristics of E coli bacteremia.

Escherichia coli O25b-ST131 and O16-ST131 causing urinary tract infection in women in Changsha, China: molecular epidemiology and clinical characteristics.

OBJECTIVE: This study aimed to investigate the prevalence of Escherichia coli ST131 and molecularly characterize the O25b-ST131 and O16-ST131 subgroups among urinary tract infection (UTI) E. coli isolates from women in central China. We also assessed the clinical characteristics and outcomes of infections caused by E. coli ST131. METHODS: Between January 2014 and December 2015, a total of 216 consecutive, non-repetitive E. coli isolates were recovered from UTI urine samples from women in Changsha, China. All isolates were analyzed for phylogenetic groups, antimicrobial resistance and virulence genotypes. ST131 clonal groups were identified using PCR and characterized using O serotyping, CTX-M genotypes, fimH, gyrA, and parC alleles, fluoroquinolone resistance genes and pulsed-field gel electrophoresis (PFGE). Clinical data were obtained from medical records. RESULTS: Overall, 41 (19.0%) of 216 E. coli isolates were identified to contain ST131 strains, among which 27 were O25b-ST131 strains and 14 were O16-ST131 strains. The clinical characteristics and outcomes of the ST131 group did not differ significantly from those of the non-ST131 group, except for the presence of urinary stones (43.9% vs 27.4%, P=0.039). Ciprofloxacin resistance was found to be significantly higher in O25b-ST131 isolates than O16-ST131 isolates (96.3% vs 14.3%, P<0.001). The majority of O25b-ST131 isolates belonged to fimH30 (92.6%), followed by fimH41 (3.7%) and fimH27 (3.7%). O25b-H30 and O25b-H41 isolates were resistant to ciprofloxacin, and possessed gyrA1AB/parC1aAB combination. All of the O16-S131 isolates were found to belong to fimH41, and of which, two of the ciprofloxacin-resistant strains harbored gyrA1AB/parC3A combination. Three PFGE clusters, consisting of 38 (92.7%) isolates, with more than 70% similarity were identified. CONCLUSION: The O25b and O16 sub-lineages have emerged as an important group of E. coli ST131 in UTI isolates from women in China. UTI patients with a history of urinary stones may need to be particularly vigilant against ST131 infection.

Epidemiology and molecular characterization of mcr-1 in Escherichia coli recovered from patients with bloodstream infections in Changsha, central China.

OBJECTIVES: The main aim of this study was to investigate the prevalence and molecular characteristics of the mcr-1 gene in Escherichia coli isolates obtained from all patients with bloodstream infections over a year in a Chinese teaching hospital. We also assessed the susceptibility profiles of the mcr-1-positive strains and prognostic impact of this gene on the patients. METHODS: A total of 144 consecutive, non-repetitive E. coli isolates causing bloodstream infections were collected at a teaching hospital in Changsha, China from January to December 2016. The presence of the mcr-1 gene was assessed by PCR. All mcr-1-positive E coli isolates were characterized by antimicrobial susceptibility testing, multilocus sequence typing (MLST), a conjugation experiment, and plasmid replicon typing. Clinical data were obtained from medical records. RESULTS: The mcr-1 gene was detected in three (2.1%) of the 144 E. coli isolates. The three mcr-1-positive E. coli isolates were resistant to colistin. All three isolates showed a lower resistance to other classes of antibacterials, with all three being susceptible to carbapenems. The MLST results indicated that the three E. coli isolates were assigned to three different sequence types: ST457, ST101, and ST1413, respectively. The conjugation experiment showed that the mcr-1 gene was successfully transferred to the recipient (E. coli EC600) from two isolates, one of which possessed IncI1 replicons and the other of which carried IncHI2 and IncN replicons. The patients with bloodstream infections caused by mcr-1-positive isolates had severe underlying diseases and were cured after antibacterial treatment. CONCLUSION: The prevalence of the mcr-1 gene in patients with E. coli bloodstream infection was 2.1% in Changsha, China. The mcr-1-positive E. coli isolates had varied susceptibility profiles, although all three were susceptible to carbapenems. This therapeutic window is crucial given the risk of rapid deterioration in high-incidence areas worldwide.

Clinical characteristics and risk factors for shock and death from E. coli bacteremia in pediatric hematological patients.

INTRODUCTION: The aim of our study was to evaluate the epidemiology, clinical features and risk factors for shock and mortality from Escherichia coli bacteremia among children and adolescents with hematological disorders. METHODOLOGY: A retrospective observational study of E. coli bacteremia in the hematology department at Xiangya Hospital from January 2013 to June 2018 was conducted. Clinical characteristics, laboratory results and antimicrobial susceptibility were analysed. Risk factors for shock and mortality were also investigated. RESULTS: Of the 45 strains of E. coli, 73.3% were multidrug-resistant (MDR). Septic shock was observed in 51.1% of patients, and the 30-day all-cause mortality was 22.2%. The risk factors associated with shock were an elevated red blood cell distribution (RDW) value when bloodstream infections (BSIs) occurred (> 15%, OR, 6.840; 95% CI, 1.571 - 29.788) and a lower WBC count (< 300/mm3, OR, 6.761; 95% CI, 1.383 - 33.044). Multivariate analysis showed that only an elevated D-dimer level (> 0.5 mg/L, OR 12.250, 95% CI 1.268 - 118.361) was a risk factor for 30-day mortality. Furthermore, we observed decreases for RDW changes at two time points (neutropenia and BSIs occurred) in the non-shock group and survival group. CONCLUSIONS: MDR infections from E. coli bacteremia were common in pediatric hematological patients. In our setting, the laboratory results may serve as a clue for physicians to distinguish patients at higher risk for shock and mortality. Furthermore, RDW could be used as a biomarker to elucidate potential disorders in hematological patients.

Fecal Carriage and Epidemiology of Carbapenem-Resistant Enterobacteriaceae Among Hospitalized Patients in a University Hospital.

PURPOSE: To determine the prevalence and epidemiology of fecal carriage of carbapenem-resistant Enterobacteriaceae (CRE), antimicrobial susceptibility, carbapenemase phenotype/genotype, and the colistin-resistance gene mcr-1 in a university hospital in China. METHODS: A comprehensive study of the fecal carriage of CRE in 704 patients was performed. Multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) were applied to elucidate the molecular epidemiology of the isolates. RESULTS: In total, 60 CRE were detected in the 704 stool samples (8.5%), including 42 Klebsiella pneumoniae, 7 Escherichia coli, 3 Citrobacter freundii, 3 Klebsiella oxytoca, 3 Enterobacter cloacae, 1 Enterobacter aerogenes, and 1 Raoultella planticola. Fifty-five CRE isolates were positive for the carbapenemase phenotype, of which 39 were Klebsiella pneumoniae carbapenemase (KPC) producers. Thirty KPC-producing K. pneumoniae sequence type (ST) 11 isolates were identified and 28 were grouped into one cluster with a similarity of ≥85%, of predominately intensive care unit (ICU) strains. Three KPC-producing ST1889 strains were isolated from the pediatric ward, all indistinguishable and resistant to tigecycline. All CRE were susceptible to colistin and negative for mcr-1. CONCLUSION: This study revealed a predominant fecal carriage of the KPC-producing K. pneumoniae ST11 clone, with several indistinguishable strain clusters, and the emergence of ST1889 in a Chinese university hospital. This evidence of cross-infection supports the urgent need for the implementation of infection control measures to prevent CRE dissemination.

Characterization of clinical extensively drug resistant Pseudomonas aeruginosa from a Chinese teaching hospital.

INTRODUCTION: Pseudomonas aeruginosa, an important opportunistic pathogen, carries multiple virulence factors which contribute to its adaptation and pathogenicity. The goal of this study was to characterize the virulence factors among extensively drug-resistant P. aeruginosa. METHODOLOGY: In this study, 63 non-duplicated extensively drug-resistant P. aeruginosa clinical isolates were collected from December 2013 to July 2015. Polymerase chain reaction (PCR) was used to analyze the homogeneity and the type III secretion system. Microtiter plate method was performed to evaluate the ability to form biofilms associated to twitching and swimming motilities. RESULTS: High percentage (96.8%) of isolates was sensitive to polymyxin B, while the resistance rate to other antibiotics (amikacin, aztreonam, ceftazidime, ciprofloxacin, gentamicin, imipenem, levofloxacin, meropenem, piperacillin-tazobactam) ranged from 80.9% to 100%. Enterobacterial repetitive intergenic consensus-PCR detected seven major groups with minimal genetic variation. All the isolates carried exoT gene, 96.8% carried exoY, 69.8% carried exoS, and 31.7% carried exoU gene. Biofilm formation was confirmed in all strains, out of which 41.3% formed strong biofilm. Motilities analysis showed heterogeneous diameters ranging from 6.02 to 26.09 mm for swimming and from 7.60 to 23.34 mm for twitching motilities. CONCLUSIONS: Our findings revealed that the clinical P. aeruginosa isolates tested are the major invasive types in nature and multiple virulence factors were commonly carried in the extensively drug-resistant strains.

[Emergence of methicillin-resistant Staphylococcus aureus SCCmec type IV/V epidemic clones in a large teaching hospital in China].

OBJECTIVE: To investigate the staphylococcal cassette chromosome mec (SCCmec) genotype and molecular epidemiological characteristics of healthcare-associated methicillin-resistant Staphylococcus aureus (HA-MRSA) in a large teaching hospital in China. METHDOS: From January 2012 to December 2012, a total of 71 nonduplicate HA-MRSA were collected in a teaching hospital in Changsha, China. SCCmec types were determined by multiplex PCR, and Panton-Valentine leukocidin (PVL) gene was detected by PCR. The homology among the tested isolates was determined using pulsed-field gel electrophoresis (PFGE). RESULTS: Of the 71 HA-MRSA isolates, 49 (69.0%) carried SCCmec III, 10 (14.1%) carried SCCmec IV, 3 (4.2%) carried SCCmec V and 3 (4.2%) carried SCCmec II; the remaining 6 isolates were not typeable by PCR. Compared with patients having SCCmec I/II/III MRSA infections, those with SCCmec IV/V MRSA infections had a significantly younger age and a similar duration of hospital stay before the first MRSA-positive culture and total hospital stay. PVL genes were strongly associated with SCCmec type IV/V MRSA infections. HA-SCCmec IV/V MRSA strains showed a greater susceptibility to rifampicin, gentamicin, levofloxacin, ciprofloxacin, and tetracycline than HA-SCCmec I/II/III MRSA strains. The 13 HA-SCCmec IV/V MRSA isolates formed one large group at the 55% similarity level. Three PFGE clusters with a similarity index of 85% or more were identified, and unique PFGE profiles were observed in 4 isolates. CONCLUSION: This is the first report of HA-MRSA isolates carrying SCCmec V in Chinese hospitals. SCCmec types IV and V MRSA clones have emerged in Chinese hospitals, which urges more rigorous surveillance of their spread in healthcare facilities in China.

[AdeABC efflux pump and resistance of Acinetobacter baumannii against carbapenem].

OBJECTIVE: To investigate relationship between AdeABC efflux pump and resistance of Acinetobacter baumannii against carbapenem.
 Methods: Carbapenem-resistant strains were acquired from multistep selection resistance test by meropenem in vitro. The quantitation test for sensitivities of strains before and after induction was determined by the E-test, and carbonylcyanide-m-chlorophenylhydrazone (CCCP) inhibition test was used to screen efflux pump. PCR, sequencing analysis, or real-time PCR was used to analyze the changes of regulatory genes adeR and adeS of the AdeABC efflux pump system, or expressions of adeA, adeB, adeR, and adeS in the strains before and after induction, respectively.
 Results: The minimal inhibitory concentrations (MICs) of meropenem were at 0.38 µg/mL and 0.25 µg/mL in parental sensitive strain S25595 and S7257, respectively, and the MICs of meropenem for both S25595 and S7257 after induction were more than 32 µg/mL. Compared with parental sensitive strains, the expression level of adeA, adeB, adeR, and adeS mRNA were elevated from 2.45 to 9.44 times, but there were no gene mutations or insertion sequences in the regulatory gene adeS and adeR.
 Conclusion: High expression of the AdeABC efflux pump system in Acinetobacter baumannii is closely associated with meropenem resistance. The upregulation of adeA and adeB expression is not due to gene mutations in the regulatory gene adeS and adeR and other mechanisms might account for it.

An Outbreak of Infections Caused by a Klebsiella pneumoniae ST11 Clone Coproducing Klebsiella pneumoniae Carbapenemase-2 and RmtB in a Chinese Teaching Hospital.

BACKGROUND: Klebsiella pneumoniae carbapenemase (KPC)-producing K. pneumoniae bacteria, which cause serious disease outbreaks worldwide, was rarely detected in Xiangya Hospital, prior to an outbreak that occurred from August 4, 2014, to March 17, 2015. The aim of this study was to analyze the epidemiology and molecular characteristics of the K. pneumoniae strains isolated during the outbreak. METHODS: Nonduplicate carbapenem-resistant K. pneumoniae isolates were screened for blaKPC-2and multiple other resistance determinants using polymerase chain reaction. Subsequent studies included pulsed-field gel electrophoresis (PFGE), multilocus sequence typing, analysis of plasmids, and genetic organization of blaKPC-2locus. RESULTS: Seventeen blaKPC-2-positive K. pneumoniae were identified. A wide range of resistant determinants was detected. Most isolates (88.2%) coharbored blaKPC-2and rmtB in addition to other resistance genes, including blaSHV-1, blaTEM-1, and aac(3)-IIa. The blaKPC-2and rmtB genes were located on the conjugative IncFIB-type plasmid. Genetic organization of blaKPC-2locusin most strains was consistent with that of the plasmid pKP048. Four types (A1, A2, A3, and B) were detected by PFGE, and Type A1, an ST11, was the predominant PFGE type. A novel K. pneumoniae sequence type (ST1883) related to ST11 was discovered. CONCLUSIONS: These isolates in our study appeared to be clonal and ST11 K. pneumoniae was the predominant clone attributed to the outbreak. Coharbing of blaKPC-2and rmtB, which were located on a transferable plasmid, in clinical K. pneumoniae isolates may lead to the emergence of a new pattern of drug resistance.

Characterization of clinical extensively drug-resistant Pseudomonas aeruginosa in the Hunan province of China.

BACKGROUND: Pseudomonas aeruginosa strains that are classed as extensively drug resistant (XDR-PA) are resistant to all antibiotics except for one or two classes and are frequently the cause of hard-to-treat infections worldwide. Our study aimed to characterize clinical XDR-PA isolates recovered during 2011-2012 at nine hospitals in the Hunan province of China. METHODS: Thirty-seven non-repetitive XDR-PA strains from 37 patients were investigated for genes encoding antimicrobial resistance determinants, efflux pumps, outer membrane proteins, and movable genetic elements using polymerase chain reaction (PCR). The expression of genes encoding the efflux pump component MexA and the outer membrane protein OprD was measured using real-time PCR. In addition, clonal relatedness of these XDR-PA isolates was analyzed by pulsed-field gel electrophoresis (PFGE). RESULTS: Various genes encoding antimicrobial resistance determinants were found in all isolates. In particular, the bla TEM-1, bla CARB, armA, bla IMP-4, bla VIM-2, and rmtB, were found in 100, 37.8, 22, 22, 19 and 5 % of the isolates, respectively. Remarkably, two isolates coharbored bla IMP-4, bla VIM-2, and armA. In all 37 antibiotic-resistant strains, the relative expression of oprD was decreased while mexA was increased compared to the expression of these genes in antibiotic-susceptible P. aeruginosa strains. All of the XDR-PA isolates harbored class I integrons as well as multiple other mobile genetic elements, such as tnpU, tnp513, tnpA (Tn21), and merA. A high genotypic diversity among the strains was detected by PFGE. CONCLUSIONS: Multiple antibiotic-resistance mechanisms contributed to the drug resistance of the XDR-PA isolates investigated in this study. Thus, the XDR-PA isolates in this area were not clonally related. Instead, multiple types of movable genetic elements were coharbored within each XDR-PA isolate, which may have aided the rapid development of these XDR-PA strains. This is the first report of XDR-PA strains that coharbor bla IMP-4, bla VIM-2, and armA.

Detection of carbapenemase-producing gram-negative bacteria using a simplified Carba NP test.

A simplified Carba NP test (CNPt), the CNPt-direct, was evaluated for carbapenemase detection using 272 gram-negative bacteria. Compared to the Modified Hodge test, the CNPt-direct had higher sensitivity (98.2% vs. 71.6%). The simplified Carba NP is an accurate, cheap and simple test for rapid carbapenemase detection.