Effect of ceftazidime-avibactam combined with different antimicrobials against carbapenem-resistant Klebsiella pneumoniae.

This study aimed to assess the in vitro efficacy of ceftazidime-avibactam (CZA) in combination with various antimicrobial agents against carbapenem-resistant Klebsiella pneumoniae (CRKP). We selected 59 clinical CRKP isolates containing distinct drug resistance mechanisms. The minimum inhibitory concentrations (MICs) of meropenem (MEM), colistin (COL), eravacycline (ERA), amikacin (AK), fosfomycin (FOS), and aztreonam (ATM), both individually and in combination with CZA, were tested using the checkerboard method. The interactions of antimicrobial agent combinations were assessed by fractional inhibitory concentration index (FICI) and susceptible breakpoint index (SBPI). The time-kill curve assay was employed to dynamically evaluate the effects of these drugs alone and in combination format. In the checkerboard assay, the combination of CZA+MEM showed the highest level of synergistic effect against both KPC-producing and carbapenemase-non-producing isolates, with synergy rates of 91.3% and 100%, respectively. Following closely was the combination of FOS+CZA . For metallo-beta-lactamases (MBLs) producing strains, ATM+CZA displayed complete synergy, while the combination of MEM+CZA showed a synergy rate of only 57.14% for NDM-producing strains and 91.67% for IMP-producing strains. In the time-kill assay, MEM+CZA also demonstrated significant synergistic effects against the two KPC-2-producing isolates (Y070 and L70), the two carbapenemase-non-producing isolates (Y083 and L093), and the NDM-1-producing strain L13, with reductions in log10 CFU/mL exceeding 10 compared to the control. Against the IMP-producing strain Y047, ATM+CZA exhibited the highest synergistic effect, resulting in a log10 CFU/mL reduction of 10.43 compared to the control. The combination of CZA and MEM exhibited good synergistic effects against KPC-producing and non-enzyme-producing strains, followed by the FOS+CZA combination. Among MBL-producing strains, ATM+CZA demonstrated the most pronounced synergistic effect. However, the combinations of CZA with ERA, AK, and COL show irrelevant effects against the tested clinical isolates. IMPORTANCE: Our study confirmed the efficacy of the combination CZA+MEM against KPC-producing and non-carbapenemase-producing strains. For metalloenzyme-producing strains, CZA+ATM demonstrated the most significant synergy. Additionally, CZA exhibited a notable synergy effect when combined with FOS. These combination therapies present promising new options for the treatment of CRKP infection.

Genomic epidemiology reveals multiple mechanisms of linezolid resistance in clinical enterococci in China.

BACKGROUND: Infections caused by linezolid-resistant enterococci (LRE) are clinically difficult to treat and threaten patient health. However, there is a lack of studies on long time-span LRE strains in China. For this reason, our study comprehensively revealed the resistance mechanisms of LRE strains collected in a Chinese tertiary care hospital from 2011 to 2022. METHODS: Enterococcal strains were screened and verified after retrospective analysis of microbial data. Subsequently, 65 LRE strains (61 Enterococcus faecalis and 4 Enterococcus faecium, MIC ≥ 8 µg/ml), 1 linezolid-intermediate Enterococcus faecium (MIC = 4 µg/ml) and 1 linezolid-susceptible Enterococcus faecium (MIC = 1.5 µg/ml) were submitted for whole-genome sequencing (WGS) analysis and bioinformatics analysis. RESULTS: The optrA gene was found to be the most common linezolid resistance mechanism in our study. We identified the wild-type OptrA and various OptrA variants in 98.5% of LRE strains (61 Enterococcus faecalis and 3 Enterococcus faecium). We also found one linezolid-resistant Enterococcus faecium strain carried both optrA and cfr(D) gene, while one linezolid-resistant Enterococcus faecium only harbored the poxtA gene. Most optrA genes (55/64) were located on plasmids, with impB-fexA-optrA, impB-fexA-optrA-erm(A), fexA-optrA-erm(A), and fexA-optrA segments. A minority of optrA genes (9/64) were found on chromosomes with the Tn6674-like platform. Besides, other possible linezolid resistance-associated mechanisms (mutations in the rplC and rplD genes) were also found in 26 enterococcal strains. CONCLUSIONS: Our study suggested that multiple mechanisms of linezolid resistance exist among clinical LRE strains in China.

Synthesis of urchin-like NiCo2S4 electrode materials based on a two-step hydrothermal method for high-capacitance supercapacitors.

Transition metal sulfides have been considered as promising electrode materials for future super-capacitors due to their spinel structures and environmentally friendly properties. Among these materials, NiCo2S4 compounds exhibit high theoretical specific capacity but poor cycling performance. To address this issue, we synthesize several NiCo2S4 urchin balls. The NCS-1.5 nanospheres demonstrate a specific capacitance of 1352.2 F g-1 at a current density of 1 A g-1, and maintain high specific capacity after 10 000 charge-discharge cycles. An asymmetric capacitor assembled with the NCS-1.5 sample as the cathode and activated carbon as the anode achieve an energy density of 45.5 W h kg-1 at 2025 W kg-1. The urchin-like nanospheres also facilitate the combination with other materials, providing potential insights for the synthesis of supercapacitor electrode materials.

Enhanced Antibacterial Ability of Electrospun PCL Scaffolds Incorporating ZnO Nanowires.

The infection of implanted biomaterial scaffolds presents a major challenge. Existing therapeutic solutions, such as antibiotic treatment and silver nanoparticle-containing scaffolds are becoming increasingly impractical because of the growth of antibiotic resistance and the toxicity of silver nanoparticles. We present here a novel concept to overcome these limitations, an electrospun polycaprolactone (PCL) scaffold functionalised with zinc oxide nanowires (ZnO NWs). This study assessed the antibacterial capabilities and biocompatibility of PCL/ZnO scaffolds. The fabricated scaffolds were characterised by SEM and EDX, which showed that the ZnO NWs were successfully incorporated and distributed in the electrospun PCL scaffolds. The antibacterial properties were investigated by co-culturing PCL/ZnO scaffolds with Staphylococcus aureus. Bacterial colonisation was reduced to 51.3% compared to a PCL-only scaffold. The biocompatibility of the PCL/ZnO scaffolds was assessed by culturing them with HaCaT cells. The PCL scaffolds exhibited no changes in cell metabolic activity with the addition of the ZnO nanowires. The antibacterial and biocompatibility properties make PCL/ZnO a good choice for implanted scaffolds, and this work lays a foundation for ZnO NWs-infused PCL scaffolds in the potential clinical application of tissue engineering.

Emergence of ST463 exoU-Positive, Imipenem-Nonsusceptible Pseudomonas aeruginosa Isolates in China.

This study investigated the resistance mechanisms and the distribution and proportions of virulence genes, including exoU, in 182 imipenem-nonsusceptible Pseudomonas aeruginosa (INS-PA) strains collected from China in 2019. There was no obvious prevalent sequence type or concentrated evolutionary multilocus sequence typing (MLST) type on the INS-PA phylogenetic tree in China. All of the INS-PA isolates harbored ß-lactamases with/without other antimicrobial mechanisms, such as gross disruption of oprD and overexpression of efflux genes. Compared with exoU-negative isolates, exoU-positive isolates (25.3%, 46/182) presented higher virulence in A549 cell cytotoxicity assays. The southeast region of China had the highest proportion (52.2%, 24/46) of exoU-positive strains. The most frequent exoU-positive strains belonged to sequence type 463 (ST463) (23.9%, 11/46) and presented multiple resistance mechanisms and higher virulence in the Galleria mellonella infection model. The complex resistance mechanisms in INS-PA and the emergence of ST463 exoU-positive, multidrug-resistant P. aeruginosa strains in southeast China indicated a challenge that might lead to clinical treatment failure and higher mortality. IMPORTANCE This study investigates the resistance mechanisms and distribution and proportions of virulence genes of imipenem-nonsusceptible Pseudomonas aeruginosa (INS-PA) isolates in China in 2019. Harboring PDC and OXA-50-like genes is discovered as the most prevalent resistance mechanism in INS-PA, and the virulence of exoU-positive INS-PA isolates was significantly higher than that of exoU-negative INS-PA isolates. There was an emergence of ST463 exoU-positive INS-PA isolates in Zhejiang, China, most of which presented multidrug resistance and hypervirulence.

Rapid automated antifungal susceptibility testing system for yeasts based on growth characteristics.

Fungal pathogens are a major threat to public health, as they are becoming increasingly common and resistant to treatment, with only four classes of antifungal medicines currently available and few candidates in the clinical development pipeline. Most fungal pathogens lack rapid and sensitive diagnostic techniques, and those that exist are not widely available or affordable. In this study, we introduce a novel automated antifungal susceptibility testing system, Droplet 48, which detects the fluorescence of microdilution wells in real time and fits growth characteristics using fluorescence intensity over time. We concluded that all reportable ranges of Droplet 48 were appropriate for clinical fungal isolates in China. Reproducibility within ±2 two-fold dilutions was 100%. Considering the Sensititre YeastOne Colorimetric Broth method as a comparator method, eight antifungal agents (fluconazole, itraconazole, voriconazole, caspofungin, micafungin, anidulafungin, amphotericin B, and 5-flucytosine) showed an essential agreement of >90%, except for posaconazole (86.62%). Category agreement of four antifungal agents (fluconazole, caspofungin, micafungin, and anidulafungin) was >90%, except for voriconazole (87.93% agreement). Two Candida albicans isolates and anidulafungin showed a major discrepancy (MD) (2.60%), and no other MD or very MD agents were found. Therefore, Droplet 48 can be considered as an optional method that is more automated and can obtain results and interpretations faster than previous methods. However, the optimization of the detection performance of posaconazole and voriconazole and promotion of Droplet 48 in clinical microbiology laboratories still require further research involving more clinical isolates in the future.

Clonal Dissemination of Antifungal-Resistant Candida haemulonii, China.

Candida haemulonii, a relative of C. auris, frequently shows antifungal resistance and is transmissible. However, molecular tools for genotyping and investigating outbreaks are not yet established. We performed genome-based population analysis on 94 C. haemulonii strains, including 58 isolates from China and 36 other published strains. Phylogenetic analysis revealed that C. haemulonii can be divided into 4 clades. Clade 1 comprised strains from China and other global strains; clades 2-4 contained only isolates from China, were more recently evolved, and showed higher antifungal resistance. Four regional epidemic clusters (A, B, C, and D) were identified in China, each comprising ≥5 cases (largest intracluster pairwise single-nucleotide polymorphism differences <50 bp). Cluster A was identified in 2 hospitals located in the same city, suggesting potential intracity transmissions. Cluster D was resistant to 3 classes of antifungals. The emergence of more resistant phylogenetic clades and regional dissemination of antifungal-resistant C. haemulonii warrants further monitoring.

Antifungal susceptibility profiles and drug resistance mechanisms of clinical Candida duobushaemulonii isolates from China.

Candida duobushaemulonii, type II Candida haemulonii complex, is closely related to Candida auris and capable of causing invasive and non-invasive infections in humans. Eleven strains of C. duobushaemulonii were collected from China Hospital Invasive Fungal Surveillance Net (CHIF-NET) and identified using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF), VITEK 2 Yeast Identification Card (YST), and internal transcribed spacer (ITS) sequencing. Whole genome sequencing of C. duobushaemulonii was done to determine their genotypes. Furthermore, C. duobushaemulonii strains were tested by Sensititre YeastOne™ and Clinical and Laboratory Institute (CLSI) broth microdilution panel for antifungal susceptibility. Three C. duobushaemulonii could not be identified by VITEK 2. All 11 isolates had high minimum inhibitory concentrations (MICs) to amphotericin B more than 2 µg/ml. One isolate showed a high MIC value of ≥64 µg/ml to 5-flucytosine. All isolates were wild type (WT) for triazoles and echinocandins. FUR1 variation may result in C. duobushaemulonii with high MIC to 5-flucytosine. Candida duobushaemulonii mainly infects patients with weakened immunity, and the amphotericin B resistance of these isolates might represent a challenge to clinical treatment.

First two fungemia cases caused by Candida haemulonii var. vulnera in China with emerged antifungal resistance.

Candida haemulonii var. vulnera is a rare variant of C. haemulonii, which has been previously reported to cause human infections. Owing to the close kinship between C. haemulonii sensu stricto and C. haemulonii var. vulnera, accurate identification of C. haemulonii var. vulnera relied on DNA sequencing assay targeting, for example, rDNA internal transcribed spacer (ITS) region. In this work, two strains of C. haemulonii var. vulnera were collected from the China Hospital Invasive Fungal Surveillance Net (CHIF-NET). The identification capacity of three matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and VITEK 2 YST ID biochemical methods were evaluated against ITS sequencing. In addition, antifungal susceptibility testing was performed using Sensititre YeastOne. Moreover, we comprehensively screened drug-resistant related genes by whole-genome sequencing. The two strains were not correctly identified to species variant level using MALDI-TOF MS and YST ID cards. Both strains were resistant to amphotericin B (minimum inhibitory concentration [MIC] > 2 µg/ml). Moreover, strain F4564 and F4584 exhibited high MIC to fluconazole (>256 µg/ml) and 5-flucytosine (>64 µg/ml), respectively, which were supposed to result from key amino acid substitutions Y132F and G307A in Erg11p and V58fs and G60K substitutions in Fur1p. The rare species C. haemulonii var. vulnera has emerged in China, and such drug-resistant fungal species that can cause invasive diseases require further close attention.

In vitro activity of lactone ketolide nafithromycin (WCK 4873) against Streptococcus pneumoniae isolates enriched with macrolide-resistance phenotype collected from mainland China.

Background: Widespread MDR Streptococcus pneumoniae in China translates clinically into a substantial pneumococcal disease burden and related morbidity and mortality, particularly in the elderly and children. Nafithromycin (WCK 4873), a novel lactone ketolide class of antibiotic designed with a 3 day, once-daily regimen is highly active against resistant pneumococci and other community respiratory pathogens. It is currently in clinical development for the treatment of community-acquired bacterial pneumonia (CABP). Objectives: To determine the in vitro activity of nafithromycin against clinical S. pneumoniae isolates collected during 2015-21 from three hospitals in mainland China. Methods: A total of 920 clinical isolates (one isolate per patient), which predominantly with the macrolide- and clindamycin-resistant phenotype were included in this study. The MICs of nafithromycin and other antibiotics tested were determined using the reference broth microdilution method. Results: Clinical S. pneumoniae isolates used in this study showed high macrolide and clindamycin resistance (>95% against erythromycin and azithromycin and 80% against clindamycin) for which nafithromycin showed potent activity (MIC50/90; 0.03/0.06 mg/L) with 100% susceptibility at a proposed pharmacokinetics/pharmacodynamics (PK/PD) breakpoint of 0.25 mg/L. Among other classes of antibiotics tested, moxifloxacin also showed good activity while amoxicillin/clavulanate and ceftriaxone showed lower susceptibility. Conclusions: Nafithromycin exhibited therapeutically relevant in vitro antibacterial activity against contemporary highly resistant pneumococci collected from mainland China. This study supports the clinical development of nafithromycin for the management of CABP caused by pneumococci in China.

Antimicrobial susceptibility to polymyxin B and other comparators against Gram-negative bacteria isolated from bloodstream infections in China: Results from CARVIS-NET program.

Objective: To investigate the bacterial distribution and antimicrobial resistance profile of clinical isolates from Gram-negative bacteria bloodstream infections (GNBSI) in China. Methods: The clinical bacterial strains isolated from blood culture were collected during April 2019 to December 2021 in 21 member hospitals of China Bloodstream Gram-negative Pathogens Antimicrobial Resistance and Virulence Surveillance Network (CARVIS-NET). Antibiotic susceptibility test was conducted by broth microdilution method recommended by Clinical and Laboratory Standards Institute (CLSI, United States). WHONET 2021 and SPSS 22.0 were used to analyze data. Results: During the study period, 1939 Gram-negative bacteria were collected from 21 hospitals, among which 1,724 (88.9%) were Enterobacteriaceae, 207 (10.7%) were non-fermenting Gram-negative bacteria and 8 (0.4%) were others. The top five bacterial species were Escherichia coli (46.2%), Klebsiella pneumoniae (31.6%), Pseudomonas aeruginosa (4.9%), Acinetobacter baumannii (4.2%) and Enterobacter cloacae (3.0%). For K. pneumoniae, antibiotic resistance was mainly prevalent in hospital-associated bloodstream infections, while for A. baumannii, antibiotic resistance was mainly prevalent in community-associated bloodstream infections. It is worth mentioning that 94.1% of the 1939 Gram-negative isolates were susceptible to polymyxin B. The sensitivity of the strains involved in our investigation to polymyxin B is highly correlated with their sensitivity to colistin. Conclusion: The surveillance results in CARVIS-NET-2021 showed that the main pathogens of GNBSI in China were Enterobacteriaceae, while E. coli was the most common pathogen. The resistance rates of K. pneumonia, P. aeruginosa, A. baumannii, and E. cloacae to multiple antibiotics kept on a high level. In many cases, polymyxin B and colistin has become the last-resort agents to combat bloodstream infections caused by multidrug-resistant (MDR) Gram-negative bacteria.

Background Filtering of Clinical Metagenomic Sequencing with a Library Concentration-Normalized Model.

Metagenomic next-generation sequencing (mNGS) can accurately detect pathogens in clinical samples. However, wet-lab contamination constrains mNGS analysis and may result in erroneous interpretation of results. Many existing methods rely on large-scale observational microbiome studies and may not be applicable to clinical mNGS tests. By generation of a pretrained profile of common laboratory contaminants, we developed an mNGS noise-filtering model based on the inverse linear relationship between microbial sequencing reads and sample library concentration, named the background elimination and correction by library concentration-normalized (BECLEAN) model. Its efficacy was evaluated with bacteria- and yeast-spiked samples and 28 cerebrospinal fluid (CSF) specimens. The diagnostic accuracy, precision, sensitivity, and specificity of BECLEAN with reference to conventional methods and diagnosis were 92.9%, 86.7%, 100%, and 86.7%, respectively. BECLEAN led to a dramatic reduction of background noise without affecting the true-positive rate and thus can provide a time-saving and convenient tool in various clinical settings. IMPORTANCE Most of the existing methods to remove wet-lab contamination rely on large-scale observational microbiome studies and may not be applicable to clinical mNGS testing in individual cases. In clinical settings, only a handful of samples might be sequenced in a run. The lab-specific microbiome can complicate existing statistical approaches for removing contamination from small-scale clinical metagenomic sequencing data sets; thus, use of a preliminary lab-specific training set is necessary. Our study provides a rapid and accurate background-filtering tool for clinical metagenomic sequencing by generation of a pretrained profile of common laboratory contaminants. Notably, our work demonstrates that the inverse linear relationship between microbial sequencing reads and library concentration can serve to identify true contaminants and evaluate the relative abundance of a taxon in samples by comparing the observed microbial reads to the model-predicted value. Our findings extend the previously published research and demonstrate confirmatory results in clinical settings.

Diagnosis of Coxiella burnetii infection via metagenomic next-generation sequencing: a case report.

BACKGROUND: Coxiella burnetii, the etiologic agent of Q fever, is mainly responsible for endocardite. But there are only a few cases of Coxiella burnetii-caused wound infection have been published, because the pathogen is very difficult to isolate using conventional culture methods. CASE PRESENTATIONS: A 76-year-old man, underwent endovascular repair of ruptured left iliac aneurysm plus abdominal aortic aneurysm under general anesthesia in 2018. Left iliac fossa mass resection was performed in 2020. After operation, the wound in the left iliac fossa was repeatedly ruptured and not healing. We used the wound tissue to perform the Metagenomics next-generation sequencing (mNGS), Coxiella burnetii was detected. Sanger sequencing and serologic verification of Coxiella burnetii all showed positive results. CONCLUSIONS: This study proved that mNGS was an effective method to detect clinically unexplained infections, and showed the ability of pathogen identification with high sensitivity and accuracy.

Optimized Sequencing Adaptors Enable Rapid and Real-Time Metagenomic Identification of Pathogens during Runtime of Sequencing.

BACKGROUND: Metagenomic next-generation sequencing (mNGS) offers the promise of unbiased detection of emerging pathogens. However, in indexed sequencing, the sequential paradigm of data acquisition, demultiplexing, and analysis restrain read assignment in advance and real-time analysis, resulting in lengthy turnaround time for clinical metagenomic detection. METHODS: We described the utility of internal-index adaptors with different lengths of barcode in multiplex sequencing. The base composition for each position within these adaptors was well-balanced to ensure nucleotide diversity and optimal sequencing performance and to achieve the early assignment of reads by first sequencing the barcodes. Combined with an automated library preparation device, we delivered a rapid and real-time bioinformatics pathogen identification solution for the Illumina NextSeq platform. The diagnostic performance was evaluated by testing 153 lower respiratory tract specimens using mNGS in comparison to culture, 16S/internal transcribed spacer amplicon sequencing, and additional PCR-based tests. RESULTS: By calculating the average F1 scores of all read lengths under different threshold values, we established the optimal threshold for pathogens identification, and found that 36 bp was the optimal shortest read length for rapid mNGS analysis. Rapid detection had a negative percentage agreement and positive percentage agreement of 100% and 85.1% for bacteria and 97.4% and 80.3% for fungi, when compared to a composite standard. The rapid mNGS solution enabled accurate pathogen identification in about 9.1 to 10.1 h sample-to-answer turnaround time. CONCLUSIONS: Optimized internal index adaptors combined with a real-time analysis pipeline provide a potential tool for a first-line test in critically ill patients.

Multicenter assessment of shotgun metagenomics for pathogen detection.

BACKGROUND: Shotgun metagenomics has been used clinically for diagnosing infectious diseases. However, most technical assessments have been limited to individual sets of reference standards, experimental workflows, and laboratories. METHODS: A reference panel and performance metrics were designed and used to examine the performance of shotgun metagenomics at 17 laboratories in a coordinated collaborative study. We comprehensively assessed the reliability, key performance determinants, reproducibility, and quantitative potential. FINDINGS: Assay performance varied significantly across sites and microbial classes, with a read depth of 20 millions as a generally cost-efficient assay setting. Results of mapped reads by shotgun metagenomics could indicate relative and intra-site (but not absolute or inter-site) microbial abundance. INTERPRETATION: Assay performance was significantly impacted by the microbial type, the host context, and read depth, which emphasizes the importance of these factors when designing reference reagents and benchmarking studies. Across sites, workflows and platforms, false positive reporting and considerable site/library effects were common challenges to the assay's accuracy and quantifiability. Our study also suggested that laboratory-developed shotgun metagenomics tests for pathogen detection should aim to detect microbes at 500 CFU/mL (or copies/mL) in a clinically relevant host context (10^5 human cells/mL) within a 24h turn-around time, and with an efficient read depth of 20M. FUNDING: This work was supported by National Science and Technology Major Project of China (2018ZX10102001).

Performance Evaluation of the Gradient Diffusion Strip Method and Disk Diffusion Method for Ceftazidime-Avibactam Against Enterobacterales and Pseudomonas aeruginosa: A Dual-Center Study.

Objectives: Ceftazidime-avibactam is a novel synthetic beta-lactam + beta-lactamase inhibitor combination. We evaluated the performance of the gradient diffusion strip method and the disk diffusion method for the determination of ceftazidime-avibactam against Enterobacterales and Pseudomonas aeruginosa. Methods: Antimicrobial susceptibility testing of 302 clinical Enterobacterales and Pseudomonas aeruginosa isolates from two centers were conducted by broth microdilution (BMD), gradient diffusion strip method, and disk diffusion method for ceftazidime-avibactam. Using BMD as a gold standard, essential agreement (EA), categorical agreement (CA), major error (ME), and very major error (VME) were determined according to CLSI guidelines. CA and EA rate > 90%, ME rate < 3%, and VME rate < 1.5% were considered as acceptable criteria. Polymerase chain reaction and Sanger sequencing were performed to determine the carbapenem resistance genes of all 302 isolates. Results: A total of 302 strains were enrolled, among which 182 strains were from center 1 and 120 strains were from center 2. A percentage of 18.21% (55/302) of the enrolled isolates were resistant to ceftazidime-avibactam. The CA rates of the gradient diffusion strip method for Enterobacterales and P. aeruginosa were 100% and 98.65% (73/74), respectively, and the EA rates were 97.37% (222/228) and 98.65% (73/74), respectively. The CA rates of the disk diffusion method for Enterobacterales and P. aeruginosa were 100% and 95.95% (71/74), respectively. No VMEs were found by using the gradient diffusion strip method, while the ME rate was 0.40% (1/247). No MEs were found by using the disk diffusion method, but the VME rate was 5.45% (3/55). Therefore, all the parameters of the gradient diffusion strip method were in line with acceptable criteria. For 31 bla KPC , 33 bla NDM , 7 bla IMP , and 2 bla VIM positive isolates, both CA and EA rates were 100%; no MEs or VMEs were detected by either method. For 15 carbapenemase-non-producing resistant isolates, the CA and EA rates of the gradient diffusion strips method were 100%. Whereas the CA rate of the disk diffusion method was 80.00% (12/15), the VME rate was 20.00% (3/15). Conclusion: The gradient diffusion strip method can meet the needs of clinical microbiological laboratories for testing the susceptibility of ceftazidime-avibactam drugs. However, the VME rate > 1.5% (5.45%) by the disk diffusion method. By comparison, the performance of the gradient diffusion strip method was better than that of the disk diffusion method.

Susceptibility to Imipenem/Relebactam of Pseudomonas aeruginosa and Acinetobacter baumannii Isolates from Chinese Intra-Abdominal, Respiratory and Urinary Tract Infections: SMART 2015 to 2018.

PURPOSE: In recent years, less options are available for treating carbapenem-resistant Acinetobacter baumannii and carbapenem-resistant Pseudomonas aeruginosa. The present study investigates the susceptibility rates to imipenem/relebactam for the treatment of intra-abdominal infections (IAIs), respiratory tract infections (RTIs) and urinary tract infections (UTIs) caused by A. baumannii and P. aeruginosa in China. PATIENTS AND METHODS: A total of 1886 P. aeruginosa and 1889 A. baumannii isolates were collected in 21 centers (7 regions) as a part of the global SMART surveillance program between 2015 and 2018. Antimicrobial susceptibility testing was performed according to the Clinical and Laboratory Standards Institute (CLSI) recommendations using the broth microdilution methodology at Peking Union Medical College Hospital. RESULTS: For P. aeruginosa, overall susceptibility rates to imipenem/relebactam were 84.2% at a CLSI breakpoint of ≤2 mg/L compared to 55.7% for imipenem. Susceptibility rates of imipenem-non-susceptible P. aeruginosa to imipenem/relebactam were 64.4% and for multidrug-resistance (MDR) P. aeruginosa susceptibility rates were increased from 25.2% for imipenem to 65.8% for imipenem/relebactam. The susceptibilities of imipenem-non-susceptible and MDR P. aeruginosa strains were similarly restored by imipenem/relebactam in non-ICU and ICU wards. The rate of imipenem-non-susceptibilities A. baumannii isolates was 79.0%, whereas the MDR rate was 81.9%. Relebactam did not change the susceptibilities of imipenem-non susceptible or MDR A. baumannii isolates. CONCLUSION: Imipenem/relebactam provides a therapy option to treat infections caused by MDR or imipenem-non-susceptible P. aeruginosa but not A. baumannii infections in China.

Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) Analysis for the Identification of Pathogenic Microorganisms: A Review.

Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) has been used in the field of clinical microbiology since 2010. Compared with the traditional technique of biochemical identification, MALDI-TOF MS has many advantages, including convenience, speed, accuracy, and low cost. The accuracy and speed of identification using MALDI-TOF MS have been increasing with the development of sample preparation, database enrichment, and algorithm optimization. MALDI-TOF MS has shown promising results in identifying cultured colonies and rapidly detecting samples. MALDI-TOF MS has critical research applications for the rapid detection of highly virulent and drug-resistant pathogens. Here we present a scientific review that evaluates the performance of MALDI-TOF MS in identifying clinical pathogenic microorganisms. MALDI-TOF MS is a promising tool in identifying clinical microorganisms, although some aspects still require improvement.

High Prevalence of Extended-Spectrum Beta-Lactamases in Escherichia coli Strains Collected From Strictly Defined Community-Acquired Urinary Tract Infections in Adults in China: A Multicenter Prospective Clinical Microbiological and Molecular Study.

OBJECTIVE: The objective of the study was to investigate the antimicrobial susceptibility and extended-spectrum beta-lactamase (ESBL) positive rates of Escherichia coli from community-acquired urinary tract infections (CA-UTIs) in Chinese hospitals. MATERIALS AND METHODS: A total of 809 E. coli isolates from CA-UTIs in 10 hospitals (5 tertiary and 5 secondary hospitals) from different regions in China were collected during the period 2016-2017 according to the strict inclusion criteria. Antimicrobial susceptibility testing was carried out by standard broth microdilution method. Isolates were categorized as ESBL-positive, ESBL-negative, and ESBL-uncertain groups according to the CLSI recommended phenotypic screening method. ESBL and AmpC genes were amplified and sequenced on ESBL-positive and ESBL-uncertain isolates. RESULTS: The antimicrobial agents with susceptibility rates of greater than 95% included imipenem (99.9%), colistin (99.6%), ertapenem (98.9%), amikacin (98.3%), cefmetazole (97.9%), nitrofurantoin (96%), and fosfomycin (95.4%). However, susceptibilities to cephalosporins (varying from 58.6% to 74.9%) and levofloxacin (48.8%) were relatively low. In the phenotypic detection of ESBLs, ESBL-positive isolates made up 38.07% of E. coli strains isolated from CA-UTIs, while 2.97% were ESBL-uncertain. Antimicrobial susceptibilities of imipenem, cefmetazole, colistin, ertapenem, amikacin, and nitrofurantoin against ESBL-producing E. coli strains were greater than 90%. The percentage of ESBL-producing strains was higher in male (53.6%) than in female patients (35.2%) (p < 0.001). CTX-M-14 (31.8%) was the major CTX-M variant in the ESBL-producing E. coli, followed by CTX-M-55 (23.4%), CTX-M-15 (17.5%), and CTX-M-27 (13.3%). The prevalence of carbapenem-resistant E. coli among CA-UTI isolates was 0.25% (2/809). CONCLUSION: Our study indicated high prevalence of ESBL in E. coli strains from strictly defined community-acquired urinary tract infections in adults in China. Imipenem, colistin, ertapenem, amikacin, and nitrofurantoin were the most active antimicrobials against ESBL-positive E. coli isolates. bla CTX-M- 14 is the predominant esbl gene in ESBL-producing and ESBL-uncertain strains. Our study indicated that the use of cephalosporins and fluoroquinolone needs to be restricted for empirical treatment of CA-UTIs in China.

Carbapenemase detection by NG-Test CARBA 5-a rapid immunochromatographic assay in carbapenem-resistant Enterobacterales diagnosis.

BACKGROUND: The global spread of carbapenem-resistant Enterobacterales (CRE) represents a serious public health concern as these organisms are associated with limited treatment options, high mortality rate and rapid transmissibility. The identification of carbapenemase remains a challenge in microbiological laboratories as no single method is perfect when considering cost, carbapenemase coverage, accuracy, handling complexity and TATs together. METHODS: NG-Test CARBA 5 assay and modified carbapenem inactivation method in conjunction with EDTA carbapenem inactivation method (mCIM/eCIM) were challenged with a collection of 299 molecularly characterized CRE isolates in China in order to evaluate the performance in detecting five major carbapenemases (bla KPC, bla NDM, bla VIM, bla IMP, and bla OXA-48) among Enterobacterales. RESULTS: NG-Test CARBA 5 detected all KPC-, NDM-, VIM- and OXA-48-producing isolates perfectly with a weak false-positive signal for NDM in an IMP-4 producer, which makes the specificity for NDM decreases to 99.6%. The overall specificity/sensitivity were 99.9%/100% for NG-Test CARBA 5. mCIM/eCIM achieved high specificity of 100%/100% and sensitivity of 99.6%/97.4%, with one S. marcescens isolate harboring VIM-2 undetected. CONCLUSIONS: Both NG-Test CARBA 5 and mCIM/eCIM showed excellent results in the tested carbapenemase (bla KPC, bla NDM, bla VIM, bla IMP, and bla OXA-48) detection compared with molecular genotypic test. As every assay has its own limitations, suitable methods should be combined for the establishment of the CRE diagnostic pathways.