Extended-Spectrum ß-Lactamase-Producing Escherichia coli and Klebsiella pneumoniae: Risk Factors and Economic Burden Among Patients with Bloodstream Infections.

Introduction: Although Extended-spectrum ß-lactamase-producing Escherichia coli and Klebsiella pneumoniae (ESBL-EK) significantly contribute to bloodstream infections, their economic repercussions remain largely unquantified. Data Source and Methods: We performed a retrospective analysis of inpatients diagnosed with Escherichia coli or Klebsiella pneumoniae bacteremia in a tertiary hospital from January 2020 to December 2022 in Guangzhou, China. We employed the chi-square test to examine ESBL risk factors and utilized propensity score matching (PSM) to negate baseline confounding factors, assessing economic burden through disability-adjusted life years (DALYs), hospital costs and productivity losses. We employed mediation analysis to eliminate confounding factors and better identify ESBL sources of burden related. Results: We found 166 ESBL-EC/KP BSI patients (52.2% of the total examined 318 patients). Post-PSM analysis revealed that ESBL-producing EC/KP will reduce the effectiveness of empirical medication by 19.8%, extend the total length of hospitalization by an average of 3 days, and increase the patient's financial burden by US$2047. No significant disparity was found in overall mortality and mean DALYs between the groups. Mediation analysis showed that the link between ESBL and hospital costs is predominantly, if not entirely, influenced by the appropriateness of empirical antibiotic treatment and length of hospital stay. Conclusion: Patients with BSI due to ESBL-producing ESBL-EK incur higher costs compared to those with non-ESBL-EK BSI. This cost disparity is rooted in varying rates of effective empirical antimicrobial therapy and differences in hospital stay durations. A nuanced approach, incorporating a thorough understanding of regional epidemiological trends and judicious antibiotic use, is crucial for mitigating the financial impact on patients.

Global emergence of a hypervirulent carbapenem-resistant Escherichia coli ST410 clone.

Carbapenem-resistant Escherichia coli (CREC) ST410 has recently emerged as a major global health problem. Here, we report a shift in CREC prevalence in Chinese hospitals between 2017 and 2021 with ST410 becoming the most commonly isolated sequence type. Genomic analysis identifies a hypervirulent CREC ST410 clone, B5/H24RxC, which caused two separate outbreaks in a children's hospital. It may have emerged from the previously characterised B4/H24RxC in 2006 and has been isolated in ten other countries from 2015 to 2021. Compared with B4/H24RxC, B5/H24RxC lacks the blaOXA-181-bearing X3 plasmid, but carries a F-type plasmid containing blaNDM-5. Most of B5/H24RxC also carry a high pathogenicity island and a novel O-antigen gene cluster. We find that B5/H24RxC grew faster in vitro and is more virulent in vivo. The identification of this newly emerged but already globally disseminated hypervirulent CREC clone, highlights the ongoing evolution of ST410 towards increased resistance and virulence.

Emergence of eravacycline heteroresistance in carbapenem-resistant Acinetobacter baumannii isolates in China.

Carbapenem-resistant Acinetobacter baumannii (CRAB) is resistant to almost all antibiotics. Eravacycline, a newer treatment option, has the potential to treat CRAB infections, however, the mechanism by which CRAB isolates develop resistance to eravacycline has yet to be clarified. This study sought to investigate the features and mechanisms of eravacycline heteroresistance among CRAB clinical isolates. A total of 287 isolates were collected in China from 2020 to 2022. The minimum inhibitory concentration (MIC) of eravacycline and other clinically available agents against A. baumannii were determined using broth microdilution. The frequency of eravacycline heteroresistance was determined by population analysis profiling (PAP). Mutations and expression levels of resistance genes in heteroresistant isolates were determined by polymerase chain reaction (PCR) and quantitative real-time PCR (qRT-PCR), respectively. Antisense RNA silencing was used to validate the function of eravacycline heteroresistant candidate genes. Twenty-five eravacycline heteroresistant isolates (17.36%) were detected among 144 CRAB isolates with eravacycline MIC values ≤4 mg/L while no eravacycline heteroresistant strains were detected in carbapenem-susceptible A. baumannii (CSAB) isolates. All eravacycline heteroresistant strains contained OXA-23 carbapenemase and the predominant multilocus sequence typing (MLST) was ST208 (72%). Cross-resistance was observed between eravacycline, tigecycline, and levofloxacin in the resistant subpopulations. The addition of efflux pump inhibitors significantly reduced the eravacycline MIC in resistant subpopulations and weakened the formation of eravacycline heteroresistance in CRAB isolates. The expression levels of adeABC and adeRS were significantly higher in resistant subpopulations than in eravacycline heteroresistant parental strains (P < 0.05). An ISAba1 insertion in the adeS gene was identified in 40% (10/25) of the resistant subpopulations. Decreasing the expression of adeABC or adeRS by antisense RNA silencing significantly inhibited eravacycline heteroresistance. In conclusion, this study identified the emergence of eravacycline heteroresistance in CRAB isolates in China, which is associated with high expression of AdeABC and AdeRS.

Practice of standardization of CLSI M45 A3 antimicrobial susceptibility testing of Infrequently Isolated or Fastidious Bacteria strains isolated from blood specimens in Guangdong Province 2017-2021.

The concentration of antimicrobial agents in environments like water and food has increased rapidly, which led to a rapid increase in antimicrobial resistance levels in the environment. Monitoring of bacterial resistance levels is considered as a necessary means to control the bacterial resistance. Reference standards are critical for antimicrobial susceptibility testing. CLSI M45 A3 standard defines pathogenic microorganisms that cause infections less frequently than those covered by CLSI M02, M07, and M100 as Infrequently Isolated or Fastidious Bacteria and specifies antimicrobial susceptibility testing methods. Our study investigated the epidemiology and antimicrobial susceptibility testing data of Infrequently Isolated or Fastidious Bacteria strains isolated from blood specimens in 70 hospitals in Guangdong Province between 2017 and 2021. We defined testing methods other than those specified in CLSI M45 A3 as "Non-Standardized." The proportion of standardized antimicrobial susceptibility testing for penicillin increased significantly (Corynebacterium spp. 17.4% vs. 50.0% p < 0.05; Micrococcus spp. 50.0% vs. 77.8% p < 0.05; Abiotrophia spp. and Granulicatella spp. 21.4% vs. 90.9% p < 0.001), while for cefotaxime (Corynebacterium spp. 0.0% vs. 45.2% p < 0.05; Abiotrophia spp. and Granulicatella spp. 0.0% vs. 14.3% p = 0.515) and vancomycin increased finitely. Non-standardized methods were used for all other antimicrobials. Due to limitations in the economic and medical environment, some clinical laboratories are unable to fully comply with CLSI M45 A3 standard. We recommend that CLSI should add breakpoints for disk diffusion method to improve the standardization of antimicrobial susceptibility testing.

Adaptive Evolution Compensated for the Plasmid Fitness Costs Brought by Specific Genetic Conflicts.

New Delhi metallo-ß-lactamase (NDM)-carrying IncX3 plasmids is important in the transmission of carbapenem resistance in Escherichia coli. Fitness costs related to plasmid carriage are expected to limit gene exchange; however, the causes of these fitness costs are poorly understood. Compensatory mutations are believed to ameliorate plasmid fitness costs and enable the plasmid's wide spread, suggesting that such costs are caused by specific plasmid-host genetic conflicts. By combining conjugation tests and experimental evolution with comparative genetic analysis, we showed here that the fitness costs related to ndm/IncX3 plasmids in E. coli C600 are caused by co-mutations of multiple host chromosomal genes related to sugar metabolism and cell membrane function. Adaptive evolution revealed that mutations in genes associated with oxidative stress, nucleotide and short-chain fatty acid metabolism, and cell membranes ameliorated the costs associated with plasmid carriage. Specific genetic conflicts associated with the ndm/IncX3 plasmid in E. coli C600 involve metabolism and cell-membrane-related genes, which could be ameliorated by compensatory mutations. Collectively, our findings could explain the wide spread of IncX3 plasmids in bacterial genomes, despite their potential cost.

Extended-Spectrum ß-Lactamase Genes Traverse the Escherichia coli Populations of Intensive Care Unit Patients, Staff, and Environment.

Over a 3-month period, we monitored the population of extended-spectrum ß-lactam-resistant Escherichia coli (ESBL-EC) associated with the patients, staff, and environment of an intensive care unit (ICU) in Guangzhou, China. Thirty-four clinical isolates were obtained from the same hospital 12 months later. A total of 165 isolates were characterized and whole-genome sequenced, with 24 isolates subjected to long-read sequencing. The diverse population included representatives of 59 different sequence types (STs). ICU patient and environmental isolates were largely distinct from staff isolates and clinical isolates. We observed five instances of highly similar isolates (0 to 13 single nucleotide polymorphisms [SNPs]) being obtained from different patients or bed unit environments. ESBL resistance in this collection was largely conferred by blaCTX-M genes, which were found in 96.4% of all isolates. The contexts of blaCTX-M genes were diverse, situated in multiple chromosomal positions and in various plasmids. We identified blaCTX-M-bearing plasmid lineages that were present in multiple STs across the surveillance, staff, and clinical collections. Closer examination of ISEcp1-blaCTX-M transposition units shed light on the dynamics of their transmission, with evidence for the acquisition of chromosomal copies of blaCTX-M genes from specific plasmid lineages and for the movement of blaCTX-M-55 from a ST1193 chromosome to a small mobilizable plasmid. A carbapenem-resistant ST167 strain isolated from a patient that had been treated with meropenem and piperacillin-tazobactam contained seven copies of blaCMY-146, which appears to have been amplified by IS1. Our data revealed limited persistence and movement of ESBL-EC strains in the ICU environment, but we observed circulating plasmid lineages playing an essential and ongoing role in shaping the cephalosporin-resistance landscape in the population examined. IMPORTANCE ESBL resistance significantly impacts clinical management of E. coli infections in hospitals globally. It is important to understand the structures of ESBL-EC populations carried by hospital patients and staff, their capacity to persist in hospital environments, and the dynamics of mobile genes that drive the spread of ESBL resistance. In our 3-month study, ESBL-EC strains found in the ICU environment were strongly associated with patient carriage but distinct from strains found in staff. However, plasmid lineages carrying blaCTX-M genes were found across the ICU populations and in a collection of clinical isolates obtained 1 year later. By examining their content and contexts, we have traced the recent histories of chromosomal and plasmid-borne ISEcp1-blaCTX-M transposition units in the ICU population. This information allowed us to implicate specific plasmid lineages in the acquisition of chromosomal blaCTX-M genes, even when the plasmids were no longer present, and to detect recent transposition of blaCTX-M-55 from a chromosome to a mobilizable plasmid. Similar high-resolution approaches to the study of mobile genetic elements will be essential if the transmission routes associated with the spread of ESBL resistance are to be understood and subjected to interventions.

Antibiotic Prescription Patterns for Acute Respiratory Infections in Rural Primary Healthcare Settings in Guangdong, China: Analysis of 162,742 Outpatient Prescriptions.

Overuse and inappropriate use of antibiotics are important contributors to bacterial antimicrobial resistance (AMR), especially in ambulatory primary healthcare (PHC) settings in low- and middle-income countries. This study aimed to investigate antibiotic prescription patterns among patients with acute respiratory infections (ARIs) in rural PHC facilities in the Guangdong Province, China. A total of 444,979 outpatient prescriptions were extracted from the electronic medical record system of 35 township health centers (THCs) and 2 community health centers (CHCs) between November 2017 and October 2018. We used the chi-square test to analyze the antibiotic prescription patterns and binary logistic regression to explore patient-related factors associated with antibiotic prescriptions. Of the 162,742 ARI prescriptions, 85.57% (n = 139,259) included at least one antibiotic. Among the 139,259 prescriptions with antibiotics, 37.82% (n = 52,666) included two or more antibiotics, 55.29% (n = 76,993) included parenteral antibiotics, and 56.62% (n = 78,852) included Watch group antibiotics. The binary logistic regression indicated that (1) female patients were slightly less likely to be prescribed antibiotics than males (adjusted odds ratio (OR) = 0.954, 95% confidence interval [CI] [0.928-0.981]; p = 0.001); and (2) compared to patients aged ≤5 years, those who were 6-15 years old (adjusted OR = 1.907, 95% CI [1.840-1.978]; p < 0.001), 16-60 years old (adjusted OR = 1.849, 95% CI [1.785-1.916]; p < 0.001), and >60 years old (adjusted OR = 1.915, 95% CI [1.810-2.026]; p < 0.001) were more likely to be prescribed antibiotics. The overuse and irrational use of antibiotics in PHC settings remain major healthcare challenges in rural Guangdong. Thus, it is imperative to implement targeted antimicrobial stewardship (AMS) policies to address this problem.

Colonization with Carbapenem-Resistant Enterobacteriaceae Contributes to Unfavorable Outcomes in End-Stage Liver Disease Patients.

Carbapenem-resistant Enterobacteriaceae (CRE) are the highest priority pathogens of the World Health Organization, and their prevalence in end-stage liver disease (ESLD) patients is increasing. CRE colonization is an independent risk factor for CRE infections. We aimed to assess risk factors and explore the relationship between CRE colonization, infection, and prognosis in patients with ESLD. A total of 311 patients with ESLD were screened for CRE colonization by fecal swabs from October 2020 to January 2022. Antimicrobial susceptibility was tested using the broth microdilution method. Carbapenem resistance genes, multilocus sequence type, and capsular serotype were analyzed by polymerase chain reaction (PCR). Seventeen CRE strains were detected, among which the most common was Klebsiella pneumoniae. The CRE colonization rate was 5.5%. Artificial liver support was an independent risk factor for CRE colonization. Compared to the non-CRE colonization group, the colonization group had a higher incidence of CRE infection and a worse prognosis. Furthermore, these strains were not closely related, and all were sensitive to polymyxin and tigecycline. There was a high colonization rate in ESLD patients, and colonization strains were highly diverse. CRE colonization deserves attention in these patients, especially when treated with artificial liver support.

In silico Evolution and Comparative Genomic Analysis of IncX3 Plasmids Isolated From China Over Ten Years.

IncX3 plasmids are correlated with the dissemination and acquisition of carbapenem resistance in Enterobacteriaceae and have been prevalent in China over the last 10 years. Since the distribution characteristics of IncX3 plasmids across China as well as their evolutionary traits for 10 years remain unclear, here we conducted a retrospective literature review and in silico comparative analysis of IncX3 plasmids in publicly available IncX3 plasmid genomes. IncX3 plasmids distributed in 17 provinces or cities were extracted for analysis, which tend to be specifically associated with hospital-isolated Escherichia coli ST410 from phylogroup A. Although the backbones of IncX3 plasmids have remained highly conservative over the last 10 years, the bla NDM resistance genetic contexts on these plasmids could fall into five subtypes, among which AR_N1_I has been identified in Enterobacter cloacae174 chromosome and AR_N5_I was simultaneously located on IncF and IncA/C plasmids. This suggests that the bla NDM resistance gene environment can spread between different plasmids, between different bacterial genera, or between strains and plasmids, highlighting that it is imperative to adopt more stringent infection control measures targeting IncX3 plasmid spread.

Mutations in porin LamB contribute to ceftazidime-avibactam resistance in KPC-producing Klebsiella pneumoniae.

Ceftazidime-avibactam (CAZ-AVI) shows promising activity against carbapenem-resistant Klebsiella pneumoniae (CRKP), however, CAZ-AVI resistance have emerged recently. Mutations in KPCs, porins OmpK35 and/or OmpK36, and PBPs are known to contribute to the resistance to CAZ-AVI in CRKP. To identify novel CAZ-AVI resistance mechanism, we generated 10 CAZ-AVI-resistant strains from 14 CAZ-AVI susceptible KPC-producing K. pneumoniae (KPC-Kp) strains through in vitro multipassage resistance selection using low concentrations of CAZ-AVI. Comparative genomic analysis for the original and derived mutants identified CAZ-AVI resistance-associated mutations in KPCs, PBP3 (encoded by ftsI), and LamB, an outer membrane maltoporin. CAZ-AVI susceptible KPC-Kp strains became resistant when complemented with mutated blaKPC genes. Complementation experiments also showed that a plasmid borne copy of wild-type lamB or ftsI gene reduced the MIC value of CAZ-AVI in the induced resistant strains. In addition, blaKPC expression level increased in four of the six CAZ-AVI-resistant strains without KPC mutations, indicating a probable association between increased blaKPC expression and increased resistance in these strains. In conclusion, we here identified a novel mechanism of CAZ-AVI resistance associated with mutations in porin LamB in KPC-Kp.

Antimutagenic specificities of two plant glycosylases, oxoguanine glycosylase and formamidopyrimidine glycosylase, assayed in vivo.

The base-excision repair process protects genomes by removing and replacing altered bases in DNA. Two analogous glycosylases, oxoguanine glycosylase (OGG) and formamidopyrimidine glycosylase (FPG), can start the process by removing oxidized guanine, the most common modification that leads to misreading of DNA. Plants possess genes for both types of glycosylases. We have tested the hypothesis that the two enzymes in plants have diverged in their specificities by inserting the genes for each enzyme from Arabidopsis thaliana L. into Escherichia coli strains designed to indicate the frequencies of the six possible single-base changes. Both enzymes retain the ability to reduce the rate of GC-->TA transversion mutations. Both enzymes also reduce the frequency of two other base-change mutations, GC-->AT and AT-->TA. We do not find a divergence in the repair capabilities of the two enzymes, as measured in E. coli, although surprisingly FPG appears to increase the rate of mutations in one particular strain.

Xyn10A, a thermostable endoxylanase from Acidothermus cellulolyticus 11B.

We cloned and purified the major family 10 xylanase (Xyn10A) from Acidothermus cellulolyticus 11B. Xyn10A was active on oat spelt and birchwood xylans between 60°C and 100°C and between pH 4 and pH 8. The optimal activity was at 90°C and pH 6; specific activity and K(m) for oat spelt xylan were 350 µmol xylose produced min⁻¹ mg of protein⁻¹ and 0.53 mg ml⁻¹, respectively. Based on xylan cleavage patterns, Xyn10A is an endoxylanase, and its half-life at 90°C was approximately 1.5 h in the presence of xylan.

A systematic review of the effectiveness and safety of Chinese herbal medicine formula Gualou Xiebai Banxia (GLXBBX) decoction for the treatment of stable angina pectoris.

BACKGROUND: A growing number of studies suggest that Gualou Xiebai Banxia (GLXBBX) decoction, a well-known Chinese herbal formula, has beneficial effects on eliminating angina pectoris symptoms and improving condition of stable angina pectoris (SAP) patients. However, whether this treatment is effective and safe for SAP or not, evidence supporting the effectiveness and safety of this treatment is still incomplete. Besides, there is lack of systematic review to assess the detailed situation (including risk of bias and methodology) of current related clinical studies. This study aimed to evaluate the effectiveness and safety of GLXBBX in treating SAP. METHODS: The major databases (MEDLINE, Embase, the Cochrane Library, China National Knowledge Infrastructure (CNKI), Chinese Scientific Journals Database (VIP) Database, Chinese Biomedical Database (CBM), Chinese Biomedical Literature Service System (SinoMed), and Wanfang Database) were searched from inception to March 2019. Randomized controlled trials (RCTs) of GLXBBX alone or combined with conventional drugs against conventional drugs for SAP were identified. Two assessors reviewed each trial independently. The methodological quality of the eligible studies was evaluated according to the Cochrane Collaboration's tool for assessing risk of bias. Both the data extraction and the literature quality screening evaluation were conducted independently by 2 researchers. RESULT: Totally 17 clinical RCTs were included in this study, involving 1676 patients. Due to the high probability of bias of the included studies, it was inappropriate to undertake a meta-analysis. Thus, we only conducted a systematic review and mainly discussed the methodology and limitation of the included studies. CONCLUSION: Although the current evidence prompted that GLXBBX might benefit SAP patients in improvement of angina pectoris, ECG, and blood lipid on a certain extent, this systematic review revealed no definite conclusion about the application of GLXBBX for SAP due to the poor methodological quality, high risk of bias, and inadequate reporting on clinical data. More rigorous, multicenter, sufficient-sample, and double-blind randomized clinical trials are warranted.

New perspectives on nodule nitrogen assimilation in actinorhizal symbioses.

Nitrogen-fixing root nodules are plant organs specialised for symbiotic transfer of nitrogen and carbon between microsymbiont and host. The organisation of nitrogen assimilation, storage and transport processes is partitioned at the subcellular and tissue levels, in distinctive patterns depending on the symbiotic partners. In this review, recent advances in understanding of actinorhizal nodule nitrogen assimilation are presented. New findings indicate that Frankia within nodules of Datisca glomerata (Presl.) Baill. carries out both primary nitrogen assimilation and biosynthesis of arginine, rather than exporting ammonium. Arginine is a typical storage form of nitrogen in plant tissues, but is a novel nitrogen carrier molecule in root nodule symbioses. Thus Frankia within D. glomerata nodules exhibits considerable metabolic independence. Furthermore, nitrogen reassimilation is likely to take place in the host in the uninfected nodule cortical cells of this root nodule symbiosis, before amino acid export to host sink tissues via the xylem. The role of an augmented pericycle in carbon and nitrogen exchange in root nodules deserves further attention in actinorhizal symbiosis, and further highlights the importance of a comprehensive, structure-function approach to understanding function in root nodules. Moreover, the multiple patterns of compartmentalisation in relation to nitrogen flux within root nodules demonstrate the diversity of possible functional interactions between host and microsymbiont that have evolved in the nitrogen-fixing clade.