Genetic, virulence, and antimicrobial resistance characteristics associated with distinct morphotypes in ST11 carbapenem-resistant Klebsiella pneumoniae.

ST11 is the most common lineage among carbapenem-resistant Klebsiella pneumoniae (CRKP) infections in Asia. Diverse morphotypes resulting from genetic mutations are associated with significant differences in microbial characteristics among K. pneumoniae isolates. Here, we investigated the genetic determinants and critical characteristics associated with distinct morphotypes of ST11 CRKP. An ST11-KL47 CRKP isolate carrying a pLVPK-like virulence plasmid was isolated from a patient with a bloodstream infection; the isolate had the "mcsw" morphotype. Two distinct morphotypes ("ntrd" and "msdw") were derived from this strain during in vitro passage. Whole genome sequencing was used to identify mutations that cause the distinct morphotypes of ST11 CRKP. Transmission electron microscopy, antimicrobial susceptibility tests, growth assays, biofilm formation, virulence assays, membrane permeability assays, and RNA-seq analysis were used to investigate the specific characteristics associated with different morphotypes of ST11 CRKP. Compared with the parental mcsw morphotype, the ntrd morphotype resulted from mutation of genes involved in capsular polysaccharide biosynthesis (wza, wzc, and wbaP), a result validated by gene knockout experiments. This morphotype showed capsule deficiency and lower virulence potential, but higher biofilm production. By contrast, the msdw morphotype displayed competition deficiency and increased susceptibility to chlorhexidine and polymyxin B. Further analyses indicated that these characteristics were caused by interruption of the sigma factor gene rpoN by insertion mutations and deletion of the rpoN gene, which attenuated membrane integrity presumably by downregulating the phage shock protein operon. These data expand current understanding of genetic, virulence, and antimicrobial resistance characteristics associated with distinct morphotypes in ST11 CRKP.

Clinical and Economic Evaluation of Blood Culture Whole Process Optimisation in Critically Ill Adult Patients With Positive Blood Cultures.

OBJECTIVES: Optimising blood culture processing is important to ensure that bloodstream infections are accurately diagnosed while minimising adverse events caused by antibiotic abuse. This study aimed to evaluate the impact of optimised blood culture processes on antibiotic use, clinical outcomes and economics in intensive care unit (ICU) patients with positive blood cultures. METHODS: From March 2020 to October 2021, this microbiology laboratory implemented a series of improvement measures, including the clinical utility of Fastidious Antimicrobial Neutralization (FAN® PLUS) bottles for the BacT/Alert Virtuo blood culture system, optimisation of bottle reception, graded reports and an upgraded laboratory information system. A total of 122 ICU patients were included in the pre-optimisation group from March 2019 to February 2020, while 179 ICU patients were included in the post-optimisation group from November 2021 to October 2022. RESULTS: Compared with the pre-optimisation group, the average reporting time of identification and antimicrobial sensitivity was reduced by 16.72 hours in the optimised group. The time from admission to targeted antibiotic therapy within 24 hours after receiving both the Gram stain report and the final report were both significantly less in the post-optimisation group compared with the pre-optimisation group. The average hospitalisation time was reduced by 6.49 days, the average antimicrobial drug cost lowered by $1720.85 and the average hospitalisation cost by $9514.17 in the post-optimisation group. CONCLUSIONS: Optimising blood culture processing was associated with a significantly increased positive detection rate, a remarkable reduction in the length of hospital stay and in hospital costs for ICU patients with bloodstream infections.

Bilateral Bow Hunter Syndrome Associated with Loss of Cervical Physiological Curvature.

BACKGROUND Bow hunter syndrome is a rare disease that is often overlooked. It presents with complex and variable clinical symptoms and causes, making diagnosis and treatment challenging. This case report focuses on a young patient with bilateral bow hunter syndrome, possibly caused by the loss of cervical physiological curvature. The aim is to enhance understanding and awareness of the disease. It is important to consider the possibility of bow hunter syndrome in young patients with long-term poor neck posture and symptoms such as dizziness, nausea, vomiting, and neck rotation-related symptoms. In such cases, thorough examination of posterior circulation hemodynamics and vascular morphology is recommended. CASE REPORT A 25-year-old woman was admitted to the hospital mainly because of "dizziness for 10 hours." The dizziness was aggravated when the right side of the neck was turned and the body position changed. This was accompanied by visual rotation, nausea, and vomiting. Bow hunter syndrome was diagnosed based on the clinical symptoms and hemodynamic examination of the posterior circulation. The patient was given a cervical collar to limit excessive twisting of the neck and instructed to avoid large-angle deflection of the neck after discharge. During the 3-month follow-up, no characteristic symptoms (such as dizziness) reappeared. CONCLUSIONS Bow hunter syndrome is a rare clinical posterior circulation compression syndrome with complex etiology. This case suggests that the simple disappearance of cervical curvature may be related to the occurrence of bow hunter syndrome. The dynamic monitoring of blood flow by color Doppler ultrasound and transcranial Doppler in different head positions provides clear clues to suspected bow hunter syndrome. With the help of computed tomography angiography, the diagnosis of bow hunter syndrome may be obtained by noninvasive examination.

The additional value of 18F-FDG PET/CT imaging in guiding the treatment strategy of non-tuberculous mycobacterial patients.

OBJECTIVES: Non-tuberculous mycobacteria (NTM) infection is an increasing health problem due to delaying an effective treatment. However, there are few data on 18F-FDG PET/CT for evaluating the status of NTM patients. The aim of this study was to investigate the potential value of 18F-FDG PET/CT in guiding the treatment strategy of NTM patients. METHODS: We retrospectively analyzed the cases of 23 NTM patients who underwent 18F-FDG PET/CT. The clinical data, including immune status and severity of NTM pulmonary disease (NTM-PD), were reviewed. The metabolic parameters of 18F-FDG included maximum standardized uptake value (SUVmax), SUVmax of the most FDG-avid lesion (SUVTop), SUVTop/SUVmax of the liver (SURLiver), SUVTop/SUVmax of the blood (SURBlood), metabolic lesion volume (MLV), and total lesion glycolysis (TLG). The optimal cut-off values of these parameters were determined using receiver operating characteristic curves. RESULTS: There were 6 patients (26.09%) with localized pulmonary diseases and 17 patients (73.91%) with disseminated diseases. The NTM lesions had high or moderate 18F-FDG uptake (median SUVTop: 8.2 ± 5.7). As for immune status, the median SUVTop in immunocompromised and immunocompetent patients were 5.2 ± 2.5 and 10.0 ± 6.4, respectively, with a significant difference (P = 0.038). As for extent of lesion involvement, SURLiver and SURBlood in localized pulmonary and disseminated diseases were 1.9 ± 1.1 vs. 3.8 ± 1.6, and 2.7 ± 1.8 vs. 5.5 ± 2.6, respectively, with a significant difference (P = 0.016 and 0.026). Moreover, for disease severity, SUVmax of the lung lesion (SUVI-lung) and SUVmax of the marrow (SUVMarrow) in the severe group were 7.7 ± 4.3 and 4.4 ± 2.7, respectively, significantly higher than those in the non-severe group (4.4 ± 2.0 and 2.4 ± 0.8, respectively) (P = 0.027 and 0.036). The ROC curves showed that SUVTop, SURLiver, SURBlood, SUVI-lung, and SUVMarrow had a high sensitivity and specificity for the identification of immune status, lesion extent, and severity of disease in NTM patients. CONCLUSION: 18F-FDG PET/CT is a useful tool in the diagnosis, evaluation of disease activity, immune status, and extent of lesion involvement in NTM patients, and can contribute to planning the appropriate treatment for NTM.

Efficacy of ceftazidime-avibactam in the treatment of carbapenem-resistant Klebsiella pneumoniae infections: Focus on solid organ transplantation recipients.

INTRODUCTION: Ceftazidime-avibactam (CAZ-AVI) is a new option to treat KPC- and OXA-48 carbapenem-resistant Klebsiella pneumoniae (CRKP) infections. However, clinical evidence is limited regarding its use in treating CRKP infections, especially in solid organ transplantation (SOT) recipients. In this study, we assessed the efficacy of CAZ-AVI in treating CRKP infections in both the general population and the SOT recipients in comparison with other antibiotic regimens. METHODS: This is a single-centre retrospective cohort study of patients admitted between January 1, 2018 and June 30, 2021 with the diagnosis of CRKP infections receiving either CAZ-AVI or other regimens ≥ 72 hours and clinical outcomes were analysed. RESULTS: Of 200 patients with CRKP infections, 67 received CAZ-AVI, 133 received other regimens, and 50 were SOT recipients. In the SOT cohort, 30 patients received CAZ-AVI, and 20 received other regimens. The overall 30-day mortality was 38% in the SOT cohort. Compared with patients receiving other regimens, CAZ-AVI therapy resulted in lower 30-day mortality (23.3% vs. 60%, P = 0.014) and 90-day mortality (35.7% vs. 86.7%, P = 0.003), higher clinical cure (93.3% vs. 40%, P < 0.001) and microbiological clearance. Similar promising results of CAZ-AVI were also shown in the whole population cohort. Moreover, clinical outcomes of SOT recipients receiving CAZ-AVI were not inferior to those without SOT. CONCLUSIONS: CAZ-AVI therapy was associated with better clinical outcomes in CRKP infections in both the general population and SOT recipients. Considering the limitations of the present study, well-conducted RCTs are still warranted to confirm these findings.

In Vitro Antibacterial Activity of Ceftobiprole and Comparator Compounds against Nation-Wide Bloodstream Isolates and Different Sequence Types of MRSA.

Bloodstream infections by bacteria, especially multidrug-resistant bacteria, remain a worldwide public health concern. We evaluated the antibacterial activity of ceftobiprole and comparable drugs against different bloodstream isolates and different sequence types of methicillin-resistant Staphylococcus aureus (MRSA) in China. We found that MRSA, methicillin-susceptible Staphylococcus aureus (MSSA), and methicillin-susceptible coagulase-negative Staphylococcus (MSCNS) displayed ceftobiprole sensitivity rates of >95%, which are similar to the rates for linezolid, daptomycin, and vancomycin. Of the tested MRCNS strains, 90.4% were sensitive to ceftobiprole. The sensitivities of ST59, ST398, and ST22 MRSA to ceftobiprole were higher than that of ST239. Ceftobiprole's MIC50/90 value against Enterococcus faecalis was 0.25/2 mg/L, whereas Enterococcus faecium was completely resistant to this drug. Ceftobiprole exhibited no activity against ESBL-positive Enterobacterales, with resistance rates between 78.6% and 100%. For ESBL-negative Enterobacterales, excluding Klebsiella oxytoca, the sensitivity to ceftobiprole was comparable to that of ceftazidime, ceftriaxone, and cefepime. The MIC50/90 value of ceftobiprole against Pseudomonas aeruginosa was 2/16 mg/L, and for Acinetobacter baumannii, it was 32/>32 mg/L. Thus, ceftobiprole shows excellent antimicrobial activity against ESBL-negative Enterobacterales and Pseudomonas aeruginosa (comparable to that of ceftazidime, ceftriaxone, and cefepime); however, it is not effective against ESBL-positive Enterobacterales and Acinetobacter baumannii. These results provide important information to clinicians.

Rapid discrimination between clinical Clostridioides difficile infection and colonization by quantitative detection of TcdB toxin using a real-time cell analysis system.

Objectives: It is important to accurately discriminate between clinical Clostridioides difficile infection (CDI) and colonization (CDC) for effective antimicrobial treatment. Methods: In this study, 37 stool samples were collected from 17 CDC and 20 CDI cases, and each sample were tested in parallel through the real-time cell analysis (RTCA) system, real-time PCR assay (PCR), and enzyme-linked immunosorbent assay (ELISA). Results: RTCA-measured functional and toxical C. difficile toxin B (TcdB) concentrations in the CDI group (302.58 ± 119.15 ng/mL) were significantly higher than those in the CDC group (18.15 ± 11.81 ng/mL) (p = 0.0008). Conversely, ELISA results revealed no significant disparities in TcdB concentrations between the CDC (26.21 ± 3.57 ng/mL) and the CDI group (17.07 ± 3.10 ng/mL) (p = 0.064). PCR results indicated no significant differences in tcdB gene copies between the CDC (774.54 ± 357.89 copies/µL) and the CDI group (4,667.69 ± 3,069.87 copies/µL) (p = 0.407). Additionally, the functional and toxical TcdB concentrations secreted from C. difficile isolates were measured by the RTCA. The results from the CDC (490.00 ± 133.29 ng/mL) and the CDI group (439.82 ± 114.66 ng/mL) showed no significant difference (p = 0.448). Notably, RTCA-measured functional and toxical TcdB concentration was significantly decreased when mixed with pooled CDC samples supernatant (p = 0.030). Conclusion: This study explored the novel application of the RTCA assay in effectively discerning clinical CDI from CDC cases.

Deep learning enables contrast-robust super-resolution reconstruction in structured illumination microscopy.

Structured illumination microscopy (SIM) is a powerful technique for super-resolution (SR) image reconstruction. However, conventional SIM methods require high-contrast illumination patterns, which necessitate precision optics and highly stable light sources. To overcome these challenges, we propose a new method called contrast-robust structured illumination microscopy (CR-SIM). CR-SIM employs a deep residual neural network to enhance the quality of SIM imaging, particularly in scenarios involving low-contrast illumination stripes. The key contribution of this study is the achievement of reliable SR image reconstruction even in suboptimal illumination contrast conditions. The results of our study will benefit various scientific disciplines.

The complete mitochondrial genome of the Baishanzu horned toad Boulenophrys baishanzuensis (Anura: Megophryidae).

The mitochondrial genome (mitogenome) of Boulenophrys baishanzuensis (Anura: Megophryidae) was sequenced by the Illumina platform. The assembled circular mitogenome of B. baishanzuensis had a total length of 17,040 bp, with a GC content of 41.25%. It consisted of 13 protein-coding genes (PCGs), two rRNA genes, 22 tRNA genes, and a D-loop region. The majority of the PCGs were encoded by the H-strand, while one PCG (nad6) and eight tRNA genes (tRNA-Gln, tRNA-Ala, tRNA-Asn, tRNA-Cys, tRNA-Tyr, tRNA-Ser2, tRNA-Glu, and tRNA-Pro) were encoded in the L-strand. Phylogenetic analysis revealed that the newly sequenced species formed a clade with other Boulenophrys species, while the genus Boulenophrys itself formed a sister group with the genus Atympanophrys.

ESKAPE in China: epidemiology and characteristics of antibiotic resistance.

The escalation of antibiotic resistance and the diminishing antimicrobial pipeline have emerged as significant threats to public health. The ESKAPE pathogens - Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter spp. - were initially identified as critical multidrug-resistant bacteria, demanding urgently effective therapies. Despite the introduction of various new antibiotics and antibiotic adjuvants, such as innovative ß-lactamase inhibitors, these organisms continue to pose substantial therapeutic challenges. People's Republic of China, as a country facing a severe bacterial resistance situation, has undergone a series of changes and findings in recent years in terms of the prevalence, transmission characteristics and resistance mechanisms of antibiotic resistant bacteria. The increasing levels of population mobility have not only shaped the unique characteristics of antibiotic resistance prevalence and transmission within People's Republic of China but have also indirectly reflected global patterns of antibiotic-resistant dissemination. What's more, as a vast nation, People's Republic of China exhibits significant variations in the levels of antibiotic resistance and the prevalence characteristics of antibiotic resistant bacteria across different provinces and regions. In this review, we examine the current epidemiology and characteristics of this important group of bacterial pathogens, delving into relevant mechanisms of resistance to recently introduced antibiotics that impact their clinical utility in China.

Comparison of the inoculum effect of in vitro antibacterial activity of Imipenem/relebactam and Ceftazidime/avibactam against ESBL-, KPC- and AmpC-producing Escherichia coli and Klebsiella pneumoniae.

OBJECTIVE: To evaluate effect of inoculum size of extended-spectrum ß-Lactamase (ESBL)-producing-, AmpC-producing-, and KPC-producing Escherichia coli and Klebsiella pneumoniae on the in vitro antibacterial effects of imipenem/relebactam (IMR) and ceftazidime/avibactam (CZA). METHODS: We compared the impact of inoculum size on IMR and CZA of sixteen clinical isolates and three standard isolates through antimicrobial susceptibility tests, time-kill assays and in vitro PK/PD studies. RESULTS: When inoculum size increased from 105 to 107 CFU/mL, an inoculum effect was observed for 26.3% (5/19) and 52.6% (10/19) of IMR and CZA, respectively; time-kill assays revealed that the concentration of CZA increased from ≥ 4 × MIC to 16 × MIC to reach 99.9% killing rate against K. pneumoniae ATCC-BAA 1705 (KPC-2-, OXA-9- and SHV-182-producing) and 60,700 (SHV-27- and DHA-1-producing). While for IMR, a concentration from 1 × MIC to 4 × MIC killed 99.9% of the four strains. When the inoculum size increased to 109 CFU/mL, neither IMR nor CZA showed a detectable antibacterial effect, even at a high concentration. An in vitro PK/PD study revealed a clear bactericidal effect when IMR administered as 1.25 g q6h when inoculum size increased. CONCLUSION: An inoculum effect on CZA was observed more frequent than that on IMR. Among the ß-lactamase-producing strains, the inoculum effect was most common for SHV-producing and KPC-producing strains.

Genome and transcriptome analysis of Enterococcus faecium from intestinal colonization and Enterococcus faecium from urinary tract infection.

Introduction: Enterococcus faecium is a common pathogen responsible for urinary tract infections (UTIs) and often establishes extensive colonization within the intestinal tract. Our aim was to assess the genomic and transcriptomic differences between colonized E. faecium without UTI (only-colonization) and colonized E. faecium causing UTI (endogenous infections). Method: We investigated the correlation between fecal isolates from the same patient and UTI-causing isolates using PFGE and WGS, and classified fecal isolates into two groups: those that solely colonized and those associated with endogenous urinary tract infections. We characterized the genomes of colonization-only and endogenously infected isolates by Scoary GWAS, and the transcriptomes of the isolates at 3 h urine exposure to assess pathogen-related changes. Result: Based on PFGE and WGS, eight isolates of endogenously infected E. faecium and nine isolates of only-colonized E. faecium were characterized and carbon and nitrogen regulated metabolisms such as genes encoding the phosphotransferase (PTS) system were enriched in endogenously infected E. faecium. Transcriptome analysis revealed significant differences in gene expression in the PTS system, lysine synthesis, galactose metabolism and citrate import between endogenously infected and only-colonized E. faecium isolates, highlighting the important role of certain carbon regulatory genes in the colonization and survival of endogenously infected E. faecium. Conclusion: In only-colonized and endogenously infected isolates, we observed differential expression patterns of genes related to carbon metabolism and amino acids, suggesting that metabolic diversity is a strategy for isolates leading to endogenous infection.

A comprehensive point prevalence survey of the quality and quantity of antimicrobial use in Chinese general hospitals and clinical specialties.

Antimicrobial resistance (AMR) is a serious, worldwide public health crisis. Surveillance of antimicrobial use forms part of an essential strategy to contain AMR. We aimed to conduct a national point prevalence survey (PPS) on antimicrobial use, and to compare this data with similar international surveillance programs to provide a reference for future AMR strategy development in China. Twenty general hospitals encompassing 10,881 beds and 10,209 inpatients around the country participated the survey using a standardized protocol, at 8am of someday from October 10th to November 31st, 2019. Of the patients, 37.00% (3777/10209) received antimicrobial agents, 31.30% (1630/5208) had surgical operations, and 76.63% (1249/1630) received prophylactic antibiotic. The prevalence of antimicrobial use in medical, surgical, and intensive care units (ICU) patients was 38.84% (1712/4408), 32.07% (1670/5208), and 66.61% (395/593), respectively. Of prescriptions, 5.79% (356/6151) were made in the absence of indication. The intensity of antimicrobial use was 61.25 DDDs/100 patient days, while the intensity of use in internal medicine, surgery, and ICU were 67.79, 45.81, 124.45 DDDs/100 patient days, respectively. Only 11.62% (715/6151) of prescriptions had a reason described in the patient record. Furthermore, 8.44% (210/2487), 14.19% (424/2989), and 12% (81/675) of the prescriptions in internal medicine, surgery, and ICU had a recorded indication, respectively. The review and stop date recorded for antimicrobial therapy was 43.73% (1976/4518). Of the patients, 38.07% (1438/3777) received combination therapy. The classes of antimicrobials prescribed were limited, and the proportion of prescriptions encompassed by the top 20 antimicrobial agents was 75.06% (4617/6151). The prevalence of antimicrobial use in China is close to that of Sweden, the UK, and Canada, but lower than that in India, and higher than that in Switzerland. The data described in this report indicate that the quality of antimicrobial prescriptions requires improvement in China. Further, hospitals should implement professional interventions to improve the rational use of antimicrobials.

pH-Responsive Polymeric Micelle Dynamic Complexes for Selective Killing of Helicobacter pylori.

Helicobacter pylori, the world's most common chronic infection-causing pathogen, is responsible for causing gastric ulcers, the fourth-leading cause of cancer-related death globally in 2020. In recent years, the effectiveness of the current treatment regimen (two antibiotics and one proton pump inhibitor) has often been plagued with problems such as resistance and the undesired elimination of commensal bacteria. Herein, we report the synthesis of block and random copolycarbonates, functionalized with cationic guanidinium and anionic acetate functional groups, aimed at selectively killing H. pylori in the acidic environment of the stomach, while remaining nontoxic to the commensal bacteria in the gut. The compositions of the polymers were fine-tuned so that the polymers were readily dispersed in water without any difficulty at both pH 3.0 and 7.4. The self-assembly behavior of the polymers at different pH values by dynamic light scattering showed that the random and block copolymers formed stable micelles in a simulated gastric environment (pH 3.0) while aggregated at pH 7.4. Both polymers demonstrated stronger antibacterial activity against H. pylori than the guanidinium-functionalized homopolymer without any acetate functional group at pH 3.0. The block copolymer was significantly more bactericidal at pH 3.0 across the concentrations tested, as compared to the random copolymer, while it did not show significant toxicity toward rat red blood cells (rRBCs) and HK-2 cells or bactericidal effect toward E. coli (a common gut bacterium) and nor caused aggregation of rRBCs at its effective concentration and at physiological pH of 7.4. Additionally, both the block and random copolymers were much more stable against hydrolysis at pH 3.0 than at pH 7.4. This study provides insight into the influence of both polymer architecture and dynamic assembly on the bioactivities of antimicrobial polymers, where the disassembly of coacervates into narrowly dispersed micelles at pH 3 make them potent antimicrobials aided by the protonated carboxylic acid block.

Clinical and microbiological characteristics of carbapenem-resistant Enterobacteriaceae causing post-operative central nervous system infections in China.

OBJECTIVES: Postoperative central nervous system infections (PCNSIs) caused by carbapenem-resistant Enterobacteriaceae (CRE) frequently result in unfavourable outcomes. However, CRE PCNSIs have not been well described from a clinical and microbiological perspective. METHODS: A total of 254 PCNSIs cases were included (January 2017 through June 2020), and clinical features were compared based on pathogenic classification. Cox regression analysis was performed to assess risk factors for mortality. Antibiotic susceptibility testing and whole genome sequencing were conducted on CRE isolates preserved. MLST, cgMLST, resistance genes and virulence genes were further analysed. RESULTS: Among 254 PCNSI cases, 15.4% were caused by Enterobacteriaceae including 28 cases by CRE. The 28-day mortality rates for CRE, CSE and non-Enterobacteriaceae PCNSIs were 50.0%, 27.3%, and 7.4%, respectively. 42.9% (12/28) of the CRE PCNSIs patients achieved clinical cure, with 25.0% achieved microbiological clearance. ST11-KL64 carrying blaKPC-2 was dominant in CRE (17/23, 73.9%), and the 28-day mortality rate of its infection was 58.5%. Most CRKP carried rampA/rampA2 genes (17/23, 73.9%). CONCLUSION: ST11-KL64 CRKP carrying blaKPC-2 dominated among CRE PCNSIs. Targeted anti-infective combination therapy based on ceftazidime/avibactam or amikacin, combined with intrathecal administration of amikacin, was found to be effective. These findings render a new insight into the clinical and microbiological landscape of CRE PCNSIs.

Comparative analysis of genomic characteristics, virulence and fitness of community-associated Staphylococcus aureus ST121 clone causing fatal diseases in China and other CA-MRSA clones.

The increasing rate of community-associated Staphylococcus aureus (CA-SA) worldwide has aroused global public concern for decades. Although ST121 clone is one of the prevalent CA-SA in China, there is still limited knowledge about it. In this study, we conducted a genomic analysis of 28 CA-SA ST121 isolates from severe bloodstream infection cases and 175 ST121 isolates from the public database. Phylogenetic analysis revealed the consistency and the complexity of global ST121 lineages, and suggested potential cross-country even cross-continental transmission of ST121 isolates. By investigating the virulence and fitness between ST121-CA-methicillin-resistant SA (CA-MRSA) and other CA-MRSA clones, we found that ST121-MRSA exhibits virulence comparable to the highly virulent USA300 clone, exceeding that of the predominant CA-MRSA lineage ST59 in China and the other American CA-MRSA clone MW2. Notably, based on analyses of virulence genes, eta, etb, edin-C and egc were only found in ST121, suggesting that the high virulence of ST121 may be attributed to the combination of these virulence factors encoded by mobile genetic elements. However, results of experiments in mice nasal and human alveolar epithelial cells showed that the colonization capacity of ST121 is much lower than that of other clones. Moreover, ST121-MRSA displayed much lower acid tolerance, suggesting that ST121-MRSA may not have such capacity to achieve the epidemiological success of other CA-MRSA clones and become the dominant lineage. Our findings expand current understanding of the epidemiology and pathogenicity of the hypervirulent ST121 clone, and highlight the importance of colonization capacity and environmental adaption in MRSA epidemiological success.

Novel SCCmec variants in clonal complex 398 and lineage-specific pseudo-SCCmec identified in ST88 MRSA from invasive bloodstream infections in China.

BACKGROUND: Methicillin resistance in Staphylococcus aureus is primarily due to the mecA gene found in highly diverse staphylococcal cassette chromosome mec (SCCmec) elements, with an increasing number of variants being continually discovered. OBJECTIVES: To characterize two novel SCCmec variants identified in clonal complex (CC) 398 strains and lineage-specific pseudo-SCCmec elements in the ST88 clone. METHODS: WGS and comparative genomic analysis were used to elucidate the SCCmec element diversity of representative isolates. RESULTS: The non-typeable 47 kb SCCmec found in the CC398 strain SKLX55795 represents a novel subtype of XIV, showing significant differences in structural organization and genetic content within the joining regions compared with the XIV element from the prototype strain SC792. This unique subtype comprised remnants from various mobile genetic elements that encode antimicrobial resistance genes, ultimately forming a large MDR region. Genome analysis of CC398 strain SKLX61416 revealed the presence of a novel 50 kb composite SCCmec with two distinct domains, carrying the ccr gene complexes 5/8 and containing genes for the detoxification of arsenic and sulphide. Further sequence analysis disclosed that 44.23% (23/52) of ST88 strains in our collection carried a lineage-specific pseudo-SCCmec, termed ΨSCCmecST88. This ΨSCCmecST88 harboured the mec gene complex C2, along with a series of genes associated with heavy metal resistance, but lacked an approximately 28 kb region encompassing the ccr gene complex. CONCLUSIONS: Our findings provide evidence for the ongoing evolution of SCCmec elements within the CC398 and ST88 clones, underscoring the need for further surveillance to understand the biological significance of these elements.

Improving Blood Culture Quality with a Medical Staff Educational Program: A Prospective Cohort Study.

Purpose: Blood cultures (BCs) are essential laboratory tests for diagnosing blood stream infections. BC diagnostic improvement depends on several factors during the preanalytical phase outside of innovative technologies. In order to evaluate the impact of an educational program on BC quality improvement, a total of 11 hospitals across China were included from June 1st 2020 to January 31st 2021. Methods: Each hospital recruited 3 to 4 wards to participate. The project was divided into three different periods, pre-implementation (baseline), implementation (educational activities administered to the medical staff) and post-implementation (experimental group). The educational program was led by hospital microbiologists and included professional presentations, morning meetings, academic salons, seminars, posters and procedural feedback. Results: The total number of valid BC case report forms was 6299, including 2739 sets during the pre-implementation period and 3560 sets during the post-implementation period. Compared with the pre-implementation period, some indicators, such as the proportion of patients who had 2 sets or more, volume of blood cultured, and BC sets per 1000 patient days, were improved in the post-implementation period (61.2% vs 49.8%, 18.56 vs 16.09 sets, and 8.0 vs 9.0mL). While BC positivity and contamination rates did not change following the educational intervention (10.44% vs 11.97%, 1.86% vs 1.94%, respectively), the proportion of coagulase negative staphylococci-positive samples decreased in BSI patients (6.87% vs 4.28%). Conclusion: Therefore, medical staff education can improve BC quality, especially increasing volume of blood cultured as the most important variable to determine BC positivity, which may lead to improved BSI diagnosis.

Genomic Epidemiology and Antimicrobial Resistance Profiles of Clostridioides difficile from Multi-Hospitals in a City in Eastern China.

Background: Clostridioides difficile is an important pathogen causing approximately 20-30% of the cases-with antibiotic-associated diarrhea and 90% of those with Pseudomembranous enteritis. However, limited surveillance of C. difficile infections (CDI) in China is done at present, especially in terms of multi-hospital epidemiological reports. Methods: Between June 2020 and November 2020, we conducted a prospective study addressing antimicrobial susceptibility profiles and genomic epidemiology of C. difficile strains isolated from inpatients with diarrhea in seven tertiary hospitals in the same city. Results: In total, 177 strains of toxin-producing C. difficile were isolated, and the dominant toxin gene profiles were tcdA+tcdB+ (84.2%, 149/177) and tcdA-tcdB+ (15.8%, 28/177). Furthermore, 130 isolates were successfully analyzed for antimicrobial susceptibility phenotype in which the rates of resistance to clindamycin, erythromycin, levofloxacin, and moxifloxacin were higher than to other antibiotics. All strains were susceptible to metronidazole and vancomycin. Fluoroquinolone-associated mutations (such as gyrA) were the most frequently found ones in the analyzed genomes. Moreover, 24 different sequence types (STs) were identified in the 130 isolates, and the most prevalent types were ST3 (26.2%, 34/130) followed by ST54 (16.9%, 22/130) and ST2 (10%, 13/130). The so-called highly virulent strain ribotyping 027 (B1/NAP1/ST1) was not identified. In addition, we also compared single nucleotide polymorphisms (SNPs) among the isolates and carried out genomic epidemiological studies on the isolates. We found that ST3 and ST54 could cause transmission in both intra- and inter-hospital settings. Conclusion: Although it is the so-called hypervirulent epidemic strain, ribotyping 027 (ST1), was not detected. ST3 and ST54 can be transmitted through different hospitals. Therefore, it is necessary to conduct further molecular epidemiological monitoring of C. difficile and screening of patients admitted to key departments.

Mechanisms of ceftazidime/avibactam resistance in drug-naïve bacteraemic Enterobacterales strains without metallo-beta-lactamase production: Associated with ceftazidime impedance.

In order to investigate ceftazidime/avibactam (CZA) resistance characteristics and mechanisms of bacteraemic Enterobacterales strains that had not been treated previously with CZA, 9708 strains were collected from 43 hospitals in 18 provinces across China from January 2019 to June 2020. The minimum inhibitory concentration (MIC) values of CZA in 165 (1.70%) strains were ≥8/4 mg/L. Ten (6.06%) CZA-resistant strains without metallo-ß-lactamase production were obtained from the individuals without prior exposure to CZA, including six Escherichia coli isolates, three Klebsiella pneumoniae isolates and one Enterobacter cloacae isolate. Whole-genome sequencing revealed that ECB88611, ECB142593 and ECB144539 had encoded disrupted OmpF loss of function. OmpF of ECB126041 had a 2_9 MKRNILAV deletion; OmpK35 of three K. pneumoniae isolates harboured amino acid fragment deletions from positions 1 to 38; and ELB117287 had encoded disrupted OmpF. The G132D amino acid substitution of OmpC of ECB88611, ECB142593 and ECB144539, and the 134_135GD insertion of OmpK36 of three K. pneumoniae isolates were predicted to alter ceftazidime permeability. 333_334 YRIK or YRIN insertions occurred in PBP3 of six E. coli isolates. The relative expression of blaKPC-2 in KPB125108 was 4.527 ± 0.2166 times higher than the control strain, and the relative expression of acrF in six E. coli isolates was 2-3 times higher than the control strain. The addition of phenylalanine-arginine-ß-naphthylamine at 100 mg/L decreased the MIC values of CZA against nine strains significantly. In conclusion, the antimicrobial resistance mechanisms in 10 isolates included increased expression of blaKPC-2, non-functional OMPs, upregulation of efflux pump activity, and variants of PBP3. Most of these mechanisms affected the antimicrobial activity of CZA by impeding ceftazidime.