Clinical Outcomes and Bacterial Characteristics of Carbapenem-resistant Acinetobacter baumannii Among Patients From Different Global Regions.

BACKGROUND: Carbapenem-resistant Acinetobacter baumannii (CRAb) is 1 of the most problematic antimicrobial-resistant bacteria. We sought to elucidate the international epidemiology and clinical impact of CRAb. METHODS: In a prospective observational cohort study, 842 hospitalized patients with a clinical CRAb culture were enrolled at 46 hospitals in five global regions between 2017 and 2019. The primary outcome was all-cause mortality at 30 days from the index culture. The strains underwent whole-genome analysis. RESULTS: Of 842 cases, 536 (64%) represented infection. By 30 days, 128 (24%) of the infected patients died, ranging from 1 (6%) of 18 in Australia-Singapore to 54 (25%) of 216 in the United States and 24 (49%) of 49 in South-Central America, whereas 42 (14%) of non-infected patients died. Bacteremia was associated with a higher risk of death compared with other types of infection (40 [42%] of 96 vs 88 [20%] of 440). In a multivariable logistic regression analysis, bloodstream infection and higher age-adjusted Charlson comorbidity index were independently associated with 30-day mortality. Clonal group 2 (CG2) strains predominated except in South-Central America, ranging from 216 (59%) of 369 in the United States to 282 (97%) of 291 in China. Acquired carbapenemase genes were carried by 769 (91%) of the 842 isolates. CG2 strains were significantly associated with higher levels of meropenem resistance, yet non-CG2 cases were over-represented among the deaths compared with CG2 cases. CONCLUSIONS: CRAb infection types and clinical outcomes differed significantly across regions. Although CG2 strains remained predominant, non-CG2 strains were associated with higher mortality. Clinical Trials Registration. NCT03646227.

Type I BREX system defends against antibiotic-resistant plasmids in Escherichia coli.

The Bacteriophage Exclusion (BREX) system is a novel antiphage defense system identified in Bacillus cereus in 2015. The purpose of this study was to investigate the presence of the BREX system defenses against antibiotic-resistant plasmids such as blaKPC and blaNDM invasion in Escherichia coli. The BREX system was present in 5.4% (23/424) of E. coli clinical isolates and 6.5% (84/1283) of E. coli strains with completely sequenced genomes in the GenBank database. All 23 BREX-positive E. coli clinical isolates were susceptible to carbapenems, while all five isolates carrying blaKPC and 11 carrying blaNDM were BREX-negative. For E. coli strains in the GenBank database, 37 of 38 strains carrying blaKPC and 109 of 111 strains carrying blaNDM were BREX negative. The recognition site sequence of methyltransferase PglX in a clinical E. coli 3756 was 5'-CANCATC-3' using PacBio single-molecular real-time sequencing. The transformation efficiency of plasmid psgRNA-ColAori-target with the PglX recognition site was reduced by 100% compared with the plasmid without the recognition site in E. coli DH5α-pHSG398-BREX. The BREX showed lower defense efficacy against plasmid psgRNA-15Aori-target which had the same plasmid backbone but different surrounding sequences of recognition sites with psgRNA-ColAori-target. The conjugation frequency of the KPC-2 plasmid and NDM-5 plasmid in E. coli 3756-ΔBREX was higher than that in E. coli 3756 clinical isolate (1.0 × 10-6 vs 1.3 × 10-7 and 5.5 × 10-7 vs 1.7 × 10-8, respectively). This study demonstrated that the type I BREX system defends against antibiotic-resistant plasmids in E. coli.

The protective effects of ruscogenin against lipopolysaccharide-induced myocardial injury in septic mice.

ABSTRACT: Sepsis-induced myocardial dysfunction (SIMD) commonly occurs in individuals with sepsis and is a severe complication with high morbidity and mortality rates. The current study aimed to investigate the effects and potential mechanisms of the natural steroidal sapogenin ruscogenin (RUS) against lipopolysaccharide (LPS)-induced myocardial injury in septic mice. We found that RUS effectively alleviated myocardial pathological damage, normalized cardiac function, and increased survival in septic mice. RNA sequencing (RNA-seq) demonstrated that RUS administration significantly inhibited the activation of the NOD-like receptor signaling pathway in the myocardial tissues of septic mice. Subsequent experiments further confirmed that RUS suppressed myocardial inflammation and pyroptosis during sepsis. Additionally, cultured HL-1 cardiomyocytes were challenged with LPS, and we observed that RUS could protect these cells against LPS-induced cytotoxicity by suppressing inflammation and pyroptosis. Notably, both the in vivo and in vitro findings indicated that RUS inhibited NLRP3 upregulation in cardiomyocytes stimulated with LPS. As expected, knockdown of NLRP3 blocked the LPS-induced activation of inflammation and pyroptosis in HL-1 cells. Furthermore, the cardioprotective effects of RUS on HL-1 cells under LPS stimulation were abolished by the novel NLRP3 agonist BMS-986299. Taken together, our results suggest that RUS can alleviate myocardial injury during sepsis, at least in part by suppressing NLRP3-mediated inflammation and pyroptosis, highlighting the potential of this molecule as a promising candidate for SIMD therapy.

Clinical characteristics and antimicrobial therapy of healthcare-associated carbapenem-non-susceptible gram-negative bacterial meningitis: a 16-year retrospective cohort study.

OBJECTIVE: Healthcare-associated Gram-negative bacterial meningitis is a substantial clinical issue with poor outcomes, especially for neurosurgical patients. Here, we aimed to study the characteristics and treatment options of patients with healthcare-associated carbapenem-non-susceptible (Carba-NS) Gram-negative bacterial meningitis. METHODS: This observational cohort study was conducted at a teaching hospital from 2004 to 2019. The clinical characteristics of patients with meningitis with Carba-NS and carbapenem-susceptible (Carba-S) bacilli were compared, and the antimicrobial chemotherapy regimens and outcomes for Carba-NS Gram-negative bacterial meningitis were analyzed. RESULTS: A total of 505 patients were included, of whom 83.8% were post-neurosurgical patients. The most common isolates were Acinetobacter spp. and Klebsiella spp., which had meropenem-resistance rates of 50.6% and 42.5%, respectively, and showed a markedly growing carbapenem-resistance trend. Kaplan-Meier curve analysis revealed that Carba-NS Gram-negative bacilli were associated with a significantly higher in-hospital mortality rate (18.8%, 35/186) compared to the Carba-S group (7.4%, 9/122; P = 0.001). For Carba-NS Enterobacterales meningitis, aminoglycoside-based and trimethoprim-sulfamethoxazole-based regimens yielded significantly higher clinical efficacy rates than non-aminoglycoside-based and non-trimethoprim-sulfamethoxazole-based regimens (69.0% vs. 38.7%, P = 0.019 and 81.8% vs. 46.9%, P = 0.036, respectively). For Carba-NS A. baumannii complex meningitis, tetracycline-based (including doxycycline, minocycline, or tigecycline) therapy achieved a significantly higher clinical efficacy rate (62.9%, 22/35) than the non-tetracycline-based therapy group (40.4%, 19/47; P = 0.044). CONCLUSIONS: Our findings revealed that Carba-NS Gram-negative bacilli are associated with higher in-hospital mortality in patients with healthcare-associated meningitis. The combination therapies involving particular old antibiotics may improve patients' outcome. TRIAL REGISTRATION: This study was registered on the Chinese Clinical Trial Register under ChiCTR2000036572 (08/2020).

International Epidemiology of Carbapenemase-Producing Escherichia coli.

BACKGROUND: Carbapenemase-producing (CP) Escherichia coli (CP-Ec) are a global public health threat. We aimed to describe the clinical and molecular epidemiology and outcomes of patients from several countries with CP-Ec isolates obtained from a prospective cohort. METHODS: Patients with CP-Ec were enrolled from 26 hospitals in 6 countries. Clinical data were collected, and isolates underwent whole-genome sequencing. Clinical and molecular features and outcomes associated with isolates with or without metallo-ß-lactamases (MBLs) were compared. The primary outcome was desirability of outcome ranking (DOOR) at 30 days after the index culture. RESULTS: Of the 114 CP-Ec isolates in Consortium on resistance against carbapenems in Klebsiella and other Enterobacterales-2 (CRACKLE-2), 49 harbored an MBL, most commonly blaNDM-5 (38/49, 78%). Strong regional variations were noted with MBL-Ec predominantly found among patients in China (23/49). Clinically, MBL-Ec were more often from urine sources (49% vs 29%), less often met criteria for infection (39% vs 58%, P = .04), and had lower acuity of illness when compared with non-MBL-Ec. Among patients with infection, the probability of a better DOOR outcome for a randomly selected patient with MBL-Ec as compared with non-MBL-Ec was 62% (95% CI: 48.2-74.3%). Among infected patients, non-MBL-Ec had increased 30-day (26% vs 0%; P = .02) and 90-day (39% vs 0%; P = .001) mortality compared with MBL-Ec. CONCLUSIONS: Emergence of CP-Ec was observed with important geographic variations. Bacterial characteristics, clinical presentations, and outcomes differed between MBL-Ec and non-MBL-Ec. Mortality was higher among non-MBL isolates, which were more frequently isolated from blood, but these findings may be confounded by regional variations.

Penicillin and Cefotaxime Resistance of Quinolone-Resistant Neisseria meningitidis Clonal Complex 4821, Shanghai, China, 1965-2020.

Clonal complex 4821 (CC4821) Neisseria meningitidis, usually resistant to quinolones but susceptible to penicillin and third-generation cephalosporins, is increasing worldwide. To characterize the penicillin-nonsusceptible (PenNS) meningococci, we analyzed 491 meningococci and 724 commensal Neisseria isolates in Shanghai, China, during 1965-2020. The PenNS proportion increased from 0.3% in 1965-1985 to 7.0% in 2005-2014 and to 33.3% in 2015-2020. Of the 26 PenNS meningococci, 11 (42.3%) belonged to the CC4821 cluster; all possessed mutations in penicillin-binding protein 2, mostly from commensal Neisseria. Genetic analyses and transformation identified potential donors of 6 penA alleles. Three PenNS meningococci were resistant to cefotaxime, 2 within the CC4821 cluster. With 96% of the PenNS meningococci beyond the coverage of scheduled vaccination and the cefotaxime-resistant isolates all from toddlers, quinolone-resistant CC4821 has acquired penicillin and cefotaxime resistance closely related to the internationally disseminated ceftriaxone-resistant gonococcal FC428 clone, posing a greater threat especially to young children.

A Novel Anti-CRISPR AcrIE9.2 Is Associated with Dissemination of blaKPC Plasmids in Klebsiella pneumoniae Sequence Type 15.

Sequence type (ST) 15 has become an emerging clone of carbapenem-resistant Klebsiella pneumoniae in which type I-E* CRISPR-Cas usually exists, indicating that the CRISPR-Cas system may not be able to block the transfer of blaKPC plasmids. The purpose of this study was to explore the mechanisms underlying dissemination of blaKPC plasmids in K. pneumoniae ST15. The type I-E* CRISPR-Cas system was present in 98.0% of 612 nonduplicate K. pneumoniae ST15 strains (88 clinical isolates and 524 from the NCBI database). Twelve ST15 clinical isolates were completely sequenced, and self-targeted protospacers were found on blaKPC plasmids flanked by a protospacer adjacent motif (PAM) of AAT in 11 isolates. The type I-E* CRISPR-Cas system was cloned from a clinical isolate and expressed in Escherichia coli BL21(DE3). In BL21(DE3) harboring the CRISPR system, the transformation efficiency of protospacer-bearing plasmids with a PAM of AAT was reduced by 96.2% compared to the empty vector, indicating that the type I-E* CRISPR-Cas system impeded blaKPC plasmid transfer. BLAST for known anti-CRISPR (Acr) amino acid sequences uncovered a novel AcrIE9-like protein with 40.5% to 44.6% sequence identity with AcrIE9 designated AcrIE9.2, which was present in 90.1% (146 of 162) of ST15 strains carrying both blaKPC and the CRISPR-Cas system. When AcrIE9.2 was cloned and expressed in a ST15 clinical isolate, the conjugation frequency of a CRISPR-targeted blaKPC plasmid was increased from 3.96 × 10-6 to 2.01 × 10-4 compared to the AcrIE9.2 absent strain. In conclusion, AcrIE9.2 may be associated with the dissemination of blaKPC in ST15 by repressing CRISPR-Cas activity.

Insertion of ISPa1635 in ISCR1 Creates a Hybrid Promoter for blaPER-1 Resulting in Resistance to Novel ß-lactam/ß-lactamase Inhibitor Combinations and Cefiderocol.

Eleven blaPER-1-positive Pseudomonas aeruginosa clinical isolates showed variable susceptibility to ceftazidime-avibactam (CZA). The genetic contexts of blaPER-1 were identical (ISCR1-blaPER-1-gst) except for the ST697 isolate HS204 (ISCR1-ISPa1635-blaPER-1-gst). The insertion of ISPa1635 in ISCR1 upstream of blaPER-1 created a hybrid promoter, which elevated the blaPER-1 transcription level and resulted in increased resistance to CZA, ceftolozane-tazobactam, cefepime-zidebactam, and cefiderocol. Diversity in the promoter activity of blaPER-1 partially explains the variable susceptibility to CZA in PER-producing isolates.

Association between a single nucleotide polymorphism of the ALOX5 gene and susceptibility to multisystem tuberculosis in a Chinese Han population.

BACKGROUND: Host genetic single nucleotide polymorphisms can exert an influence susceptibility to tuberculosis infection. Previous investigations have demonstrated an association between the polymorphism in the ALOX5 gene and a range of diseases, encompassing not only noninfectious conditions like asthma, acute myocardial infarction, and cerebral infarction but also infections caused by various pathogens. However, the relationship between ALOX5 gene polymorphism and susceptibility to tuberculosis has received limited research attention. The ALOX5 gene encodes arachidonic acid 5-lipoxygenase(5-LO), which serves as the initiating catalyst in the generation of the inflammatory mediator leukotriene. Leukotrienes, products derived from the 5-LO pathway, are potent proinflammatory lipid mediators that assume a pivotal role in tuberculosis infections.Consequently, ALOX5 gene variants may be intricately associated with the pathogenesis of tuberculosis. In instances where the host exhibits immunocompromisation, infection with Mycobacterium tuberculosis can impact multiple systems. The involvement of multiple systems significantly augments the complexity of treatment and escalates patient mortality rates. Regrettably, the underlying mechanisms driving multisystem tuberculosis pathogenesis remain enigmatic, with clinicians paying scant attention to this aspect. Although the protein encoded by the ALOX5 gene represents a pivotal enzyme that catalyzes the metabolism of arachidonic acid into LXA4, and thereby plays a significant role in the inflammatory response during tuberculosis infection, studies investigating ALOX5 gene polymorphism and its association with susceptibility to multisystem tuberculosis in the Chinese Han population are exceptionally scarce. Therefore, the primary objective of this study is to comprehensively examine the correlation between ALOX5 gene polymorphisms and susceptibility to tuberculosis within the Chinese Han population, with particular emphasis on multisystemic tuberculosis. METHODS: A case‒control study design was employed, encompassing 382 individuals with pulmonary tuberculosis and 367 individuals with multisystemic tuberculosis as the case groups, along with 577 healthy controls.Whole blood DNA was extracted from all patients and healthy controls. Subsequently, three tag polymorphisms (rs2029253, rs7896431, rs2115819) within the ALOX5 gene were selectively identified and genotyped. RESULTS: After adjusting for age and sex, the presence of allele A at rs2029253 exhibited a pronounced association with an elevated risk of TB susceptibility when compared to the tuberculosis group and healthy control group. (ORa: 2.174, 95% CI: 1.827-2.587; Pa<0.001, respectively). Notably, the rs2029253 AG genotype and AA genotype displayed a significantly increased susceptibility to tuberculosis (ORa: 2.236, 95% CI: 1.769-2.825; Pa <0.001 and ORa: 4.577, 95% CI: 2.950-7.100; Pa <0.001, respectively) compared to the GG genotype. Moreover, in the analysis utilizing genetic models, rs2029253 also exhibited a markedly heightened susceptibility to tuberculosis in additive models, dominant models, and recessive models (Pa <0.001). Conversely, no significant association was observed between rs7896431, rs2115819, and tuberculosis. In the subgroup analysis, when comparing the pulmonary tuberculosis group with the healthy control group, we observed no significant disparities in the distribution frequencies of alleles, genotypes, and gene models (additive model, dominant model, and recessive model) for the three tag SNPs, with P-values were >0.05 after adjusting for age and sex. Additionally, we noted that the presence of allele A at rs2029253 was linked to an increased susceptibility to tuberculosis in the multisystemic tuberculosis group relative to the healthy control group (ORa: 2.292, 95% CI: 1.870-2.810; Pa<0.001). Similarly, the rs2029253 AG genotype, AA genotype, and gene models, including the additive model, dominant model, and recessive model, demonstrated a significantly elevated risk of tuberculosis susceptibility. CONCLUSIONS: The polymorphism in the ALOX5 gene is associated with susceptibility to multisystemic tuberculosis in the Chinese Han population.

Disk diffusion-based method aids in the detection of vanM-positive Enterococcus faecium with low vancomycin minimum inhibitory concentrations.

We first reported the vanM vancomycin resistance gene in enterococci in Shanghai, China in 2006 and later found it to be the predominant van gene in vancomycin-resistant enterococci (VRE). In this study, we successively collected 1292 Enterococcus faecium and Enterococcus faecalis strains from in- and outpatients at Huashan Hospital, Fudan University and found that nearly all of the isolates (1290/1292) were vancomycin-sensitive determined by the VITEK 2 system. However, using a modified macromethod-based disk diffusion test, 10 E. faecium isolates that were previously determined to be vancomycin-sensitive by the VITEK 2 system were found to have colonies in the vancomycin disk inhibition zone. Pulse-field gel electrophoresis results showed that each randomly selected colony in the inhibition zone belonged to the same clone as the original strain. All 10 isolates were later found to be vanM-positive. The disk diffusion-based method may aid in the detection of vanM-positive E. faecium with low vancomycin minimum inhibitory concentrations and prevent missing the detection of vancomycin sensitivity-variable enterococci.

Physicians' knowledge of invasive fungal disease in China.

BACKGROUND: Invasive fungal disease (IFD) is associated with high morbidity and mortality. Data are lacking regarding physicians' perspectives on the diagnosis and management of IFD in China. OBJECTIVES: To evaluate physicians' perspectives on the diagnosis and management of IFD. METHODS: Based on current guidelines, a questionnaire was designed and administered to 294 physicians working in haematology departments, intensive care units, respiratory departments and infectious diseases departments in 18 hospitals in China. RESULTS: The total score and subsection scores for invasive candidiasis, invasive aspergillosis (IA), cryptococcosis and invasive mucormycosis (IM) were 72.0 ± 12.2 (maximum = 100), 11.1 ± 2.7 (maximum = 19), 43.0 ± 7.8 (maximum = 57), 8.1 ± 2.0 (maximum = 11) and 9.8 ± 2.3 (maximum = 13), respectively. Although the perspectives of the Chinese physicians were in good overall agreement with guideline recommendations, some knowledge gaps were identified. Specific areas in which the physicians' perspectives and guideline recommendations differed included use of the ß-D-glucan test to facilitate the diagnosis of IFD, relative utility of the serum galactomannan test and bronchoalveolar lavage fluid galactomannan test in patients with agranulocytosis, use of imaging in the diagnosis of mucormycosis, risk factors for mucormycosis, indications for initiating antifungal therapy in patients with haematological malignancies, when to start empirical therapy in mechanically ventilated patients, first-line drugs for mucormycosis and treatment courses for IA and IM. CONCLUSION: This study highlights the main areas that could be targeted by training programs to improve the knowledge of physicians treating patients with IFD in China.

Association between a single nucleotide polymorphism of the IL23R gene and tuberculosis in a Chinese Han population: a case‒control study.

BACKGROUND: Severe tuberculosis constitutes a significant menace to human safety and well-being, with a considerable mortality rate. The severity of tuberculosis can be impacted by genetic variations in host genes, particularly single nucleotide polymorphisms (SNPs). METHODS: A case‒control study was undertaken, encompassing a cohort of 1137 tuberculosis patients (558 with severe tuberculosis and 579 with mild tuberculosis), alongside 581 healthy controls within the age range of fifteen to forty-five years. Whole blood DNA was extracted from all participants, and three tag polymorphisms (rs1884444, rs7518660, rs7539625) of the IL23R gene were selectively identified and genotyped. RESULTS: No significant correlation was observed between the IL23R gene polymorphisms (rs1884444, rs7518660, and rs7539625) and tuberculosis. Upon comparing the tuberculosis group with the healthy control group, the mild tuberculosis group with the healthy control group, and the severe tuberculosis group with the healthy control group, the obtained P-values were> 0.05. However, in the comparison between severe tuberculosis and mild tuberculosis, the presence of rs1884444 G alleles exhibited a significantly increased risk of severe tuberculosis after adjusting for age and sex (ORa: 1.199, 95% CI: 1.009-1.424; Pa=0.039, respectively). In subgroup analysis, after accounting for confounding factors, including age and sex, rs1884444 G alleles continued to demonstrate a significantly heightened risk of severe tuberculosis. Nonetheless, the comparison between the multisystemic tuberculosis group and the mild tuberculosis group was no significant difference. Notably, rs1884444 of the IL23R gene exhibited a noteworthy association with the risk of severe tuberculosis in the comparison between severe tuberculosis and mild tuberculosis before and after adjusting for age and sex (ORa: 1.301, 95% CI: 1.030-1.643; Pa=0.027, respectively). Furthermore, the presence of the rs1884444 G allele exhibited a significantly increased risk of severe tuberculosis after adjusting for age and sex in the comparison between tuberculous meningitis and mild tuberculosis (ORa: 1.646, 95% CI: 1.100-2.461; Pa=0.015, respectively). CONCLUSIONS: The present study suggests that there is no significant association between IL23R gene polymorphism and tuberculosis susceptibility in the Chinese Han population. However, it does indicate a potential link between IL23R polymorphism and an increased risk of developing severe tuberculosis.

Serogroup Y Clonal Complex 23 Meningococcus in China Acquiring Penicillin Resistance from Commensal Neisseria lactamica Species.

Invasive meningococcal disease (IMD) due to serogroup Y Neisseria meningitidis (NmY) is rare in China; recently, an invasive NmY isolate, Nm512, was discovered in Shanghai with decreased susceptibility to penicillin (PenNS). Here, we investigated the epidemiology of NmY isolates in Shanghai and explored the potential commensal Neisseria lactamica donor of the PenNS NmY isolate. A total of 491 N. meningitidis and 724 commensal Neisseria spp. isolates were collected. Eleven NmY isolates were discovered from IMD (n = 1) and carriers (n = 10), including two PenNS isolates with five-key-mutation-harboring (F504L-A510V-I515V-H541N-I566V) penA genes. Five of the eight ST-175 complex (CC175) isolates had a genotype [Y:P1.5-1,2-2:F5-8:ST-175(CC175)] identical to that of the predominant invasive clone found in South Africa. Only one invasive NmY CC23 isolate (Nm512) was discovered; this isolate carried a novel PenNSpenA832 allele, which was identified in commensal N. lactamica isolates locally. Recombination analysis and transformation of the penA allele highlighted that N. meningitidis Nm512 may acquire resistance from its commensal donor; this was supported by the similar distribution of transformation-required DNA uptake sequence variants and the highly cognate receptor ComP between N. meningitidis and N. lactamica. In 2,309 NmY CC23 genomes from the PubMLST database, isolates with key-mutation-harboring penA genes comprised 12% and have been increasing since the 1990s, accompanied by recruitment of the blaROB-1 and/or quinolone resistance allele. Moreover, penA22 was predominant among genomes without key mutations in penA. These results strongly suggest that Nm512 is a descendant of the penA22-harboring CC23 isolate from Europe and acquired its penicillin resistance locally from commensal N. lactamica species by natural transformation.

Transmission barrier of the blaKPC plasmid mediated by type I restriction-modification systems in Escherichia coli.

BACKGROUND: Transportation of carbapenem-resistant plasmids contributes to carbapenem resistance in Gram-negative bacteria. KPC enzymes are the most clinically important enzymes among carbapenem-resistant Klebsiella pneumoniae, whereas the rate of blaKPC in Escherichia coli is low. The CRISPR-Cas system and restriction-modification system (R-M system) in bacteria defend against invading genomes. Currently, the role of the immune systems in the low rate of KPC-producing E. coli remains unclear. OBJECTIVES: We investigated the relationship between immune systems and the low detection rate of blaKPC in E. coli. METHODS: We searched for blaKPC among 1039 E. coli whole genomes available in GenBank using nucleotide BLAST. CRISPR-Cas systems and the R-M system were detected in all strains having the ST as blaKPC-positive strains. Nucleotide BLAST was used to search for protospacers on blaKPC plasmids. A conjugation assay was performed to determine whether the R-M system influences the acquisition of blaKPC plasmids by E. coli. RESULTS: ST131 was the dominant ST of KPC-producing E. coli and IncN was the main plasmid type (12/32). CRISPR-Cas systems were frequently present in E. coli carrying blaKPC. Furthermore, CRISPR-Cas systems in E. coli didn't target plasmids with blaKPC. Type I R-M systems were rare in KPC-producing E. coli, but significantly over-represented in KPC-negative strains. E. coli DH5α with hsdR deletion accepted blaKPC-carrying plasmids, whereas those with hsdR complementation impeded blaKPC-carrying plasmid conjugation. CONCLUSIONS: Horizontal transmission of blaKPC occurs among E. coli. The type I R-M system is associated with the defence against blaKPC plasmid transport into E. coli.

Metabolomics and microbiomes for discovering biomarkers of antituberculosis drugs-induced hepatotoxicity.

Anti-tuberculosis (TB) drug-induced hepatotoxicity (ATDH) was related to metabolic and microbial dysregulation, but only limited data was available about the metabolomes and microbiomes in ATDH. We aimed at detecting the metabolic and microbial signatures of ATDH. Urine samples were obtained from ATDH (n = 33) and non-ATDH control (n = 41) and analyzed by untargeted gas chromatography time-of-flight mass spectrometry (GC-TOF-MS). Metabolites were analyzed by orthogonal projections to latent structures-discriminate analysis (OPLS-DA) and pathway analysis. Eight ATDH and eight non-ATDH control were evaluated by sequencing of 16S rRNA genes, and the Clusters of Orthologous Groups of proteins (COG) database were used for function prediction. Linear discriminant analysis (LDA) effect size (LEfSe) was applied to detect the differential microbiotas between the two groups. The differential microbiotas were further validated by correlation analysis with differential metabolites. OPLS-DA analysis suggested 11 metabolites that differed ATDH from non-ATDH control. Pathway analysis demonstrated that metabolism of arginine and proline, metabolism of d-arginine and d-ornithine, glutathione glycine metabolism, galactose metabolism, niacin and nicotinamide metabolism, and glycine, serine and threonine metabolism were related to ATDH. LEfSe suggested significant differences in microbiotas between the two groups. The o_ Bacteroidales, f_Prevotellaceae, and g_Prevotella were significantly increased in ATDH. In contrast, the f_Chitinophagaceae, c_Gammaproteobacteria, and p_Proteobacteria were significantly increased in non-ATDH group. The biological functions of the sequenced microbiota in this study were related to amino acid transport and metabolism and defense mechanisms. Finally, we detected strong association between urine metabolites and specific urine bacteria (|r| > 0.8). d-glucoheptose showed a strong relationship to Symbiobacterium. Creatine (r = -0.901; P < 0.001) and diglycerol were strongly associated with Alishewanella. Metabolomics and microbiomes indicate ATDH characterized by metabolic and microbial profiles may differ from non-ATDH control.

Treatment and economic burden of mucormycosis in China: Case report review and burden estimation.

WHAT IS KNOWN AND OBJECTIVE: Mucormycosis is an opportunistic fungal infection associated with low incidence but high mortality. Few studies have shown the treatment and disease burden of mucormycosis in China. This study aims at collecting all the reported cases to describe the characteristics and treatment patterns and to assess the economic burden of mucormycosis in China. METHODS: We conducted a literature review of mucormycosis case reports in Chinese patients to summarize the characteristics and treatment patterns of the disease in China. An economic model was built to evaluate the total cost of mucormycosis per person, including direct medical cost, direct non-medical cost and indirect cost. RESULTS AND DISCUSSION: A total of 676 case reports showed that the most common type of mucormycosis was pulmonary mucormycosis (299/676, 44.2%), and rhinocerebral mucormycosis had the highest case fatality rate (122/185, 68.5%). Among those who used empiric therapies, 48.8% (231/473) did not include anti-mucor drugs; 79.8% (336/421) of the therapies include amphotericin B (AMB) or AMB-lipo after detection of mucormycetes; 98.6% (69/70) of the reported adverse events were associated with AMB and AMB-lipo. The duration of treatment ranged from 90 to 180 days; the length of stay ranged from 22 to 95 days. The average total cost per patient was 166 thousand Chinese Yuan (CNY), of which 93.1% was the direct medical cost (155 thousand CNY). WHAT IS NEW AND CONCLUSION: There are a limited number of antifungal treatment options for mucormycosis in China. This study highlights the critical need to introduce innovative and broader spectrum antifungal drugs with improved safety, better clinical efficacy, easier administration and reduced economic burden to Chinese mucormycosis patients.

Identification of qnrE3 and qnrE4, New Transferable Quinolone Resistance qnrE Family Genes Originating from Enterobacter mori and Enterobacter asburiae, Respectively.

The qnrE family was designated in 2017. To date, two qnrE alleles have been discovered that are carried by plasmids. Here, we identified a new quinolone resistance gene, qnrE3, in the chromosome of Enterobacter mori clinical isolate 08-091 in China. qnrE3 conferred decreased susceptibility to fluoroquinolones, similar to qnrE1 and qnrE2. To investigate the precise origin of qnrE1, qnrE2, and qnrE3, 79 qnrE-bearing strains producing 30 qnrE variants were retrieved from the NCBI database. Phylogenetic analysis illustrated two major clusters, QnrEEmo and QnrEEas, produced mainly by the E. mori and E. asburiae strains, respectively. Comparison of the genetic context of qnrE alleles demonstrated that qnrE3 and qnrEEas2 alleles presumably were captured by ISEcp1 and mobilized from the E. mori and E. asburiae strains to the E. xiangfangensis and Escherichia coli strains, respectively. qnrEEas2 was proposed to be named qnrE4, since it has spread to another genus. All the qnrE alleles were harbored by the Enterobacter species, except those captured by ISEcp1 and mobilized into other species of Enterobacterales. E. mori is probably the source of qnrE1 to qnrE3 alleles, and E. asburiae is the reservoir of qnrE4.

Development of a Fast Raman-Assisted Antibiotic Susceptibility Test (FRAST) for the Antibiotic Resistance Analysis of Clinical Urine and Blood Samples.

Human health is at great risk due to the spreading of antimicrobial resistance (AMR). The lengthy procedure of conventional antimicrobial susceptibility testing (AST) usually requires a few days. We developed a fast Raman-assisted antibiotic susceptibility test (FRAST), which detects single bacterial metabolic activity in the presence of antibiotics, using Raman single-cell spectroscopy. It was found that single-cell Raman spectra (SCRS) would show a clear and distinguishable Raman band at the "silent zone" (2000-2300 cm-1), due to the active incorporation of deuterium from heavy water (D2O) by antibiotic-resistant bacteria. This pilot study has compared the FRAST and the conventional AST for six clinical standard quality controls (four Gram-negative and two Gram-positive bacteria strains) in response to 38 antibiotics. In total, 3200 treatments have been carried out and approximately 64 000 SCRS have been acquired for FRAST analysis. The result showed an overall agreement of 88.0% between the FRAST and the conventional AST assay. The gram-staining classification based on the linear discriminant analysis (LDA) model of SCRS was developed, seamlessly coupling with the FRAST to further reduce the turnaround time. We applied the FRAST to real clinical analysis for nine urinary infectious samples and three sepsis samples. The results were consistent with MALDI-TOF identification and the conventional AST. Under the optimal conditions, the "sample to report" of the FRAST could be reduced to 3 h for urine samples and 21 h for sepsis samples. The FRAST provides fast and reliable susceptibility tests, which could speed up microbiological analysis for clinical practice and facilitate antibiotic stewardship.

Characterization of the novel plasmid-encoded MBL gene blaAFM-1, integrated into a blaIMP-45-bearing transposon Tn6485e in a carbapenem-resistant Pseudomonas aeruginosa clinical isolate.

OBJECTIVES: To characterize the novel subclass B1 MBL AFM-1, encoded by a blaIMP-45-bearing megaplasmid from a carbapenem-resistant Pseudomonas aeruginosa (CRPA) clinical isolate. METHODS: CRPA HS17-127 and its transconjugant were discovered to carry blaAFM-1 in our previous study. blaAFM-1 and blaNDM-1 were cloned and expressed in Escherichia coli TOP10 and P. aeruginosa PAO1, respectively, to test the resistance phenotype. Kinetic studies were performed to elucidate the biochemical characteristics of the AFM-1 enzyme. Comparative genomic analysis was applied to investigate the genetic context of blaAFM-1. RESULTS: PAO1 transconjugant TcHS17-127 exhibited carbapenem resistance with an imipenem MIC of 64 mg/L. E. coli transformants with cloned blaAFM-1 or blaNDM-1 had increased MICs of all ß-lactams tested (except aztreonam) and imipenem MICs of 4-8 mg/L. Kinetic studies showed that AFM-1 had greater catalytic efficiency against cephalosporins than carbapenems. blaAFM-1 was located on a 486 963 bp IncP-2 plasmid, pHS17-127, containing a 57.3 kb MDR Tn1403-derivative transposon, Tn6485e, which is genetically closest to the blaIMP-45-bearing Tn6485 transposon but has acquired an extra ISCR27n3-blaAFM-1 module. Multicentre surveillance of 605 P. aeruginosa clinical isolates identified three blaAFM carriers from different STs. Two of them co-carried blaAFM-1 and blaIMP-45. A BLAST search against the NCBI database showed six blaAFM carriers on various plasmids and the chromosomes of different Gram-negative species. CONCLUSIONS: The blaAFM-1 gene confers carbapenem resistance and has been captured in distinct species of non-fermenters. Co-carriage of blaAFM-1 and blaIMP-45 in an MDR transposon on a conjugative plasmid can be expected to promote further dissemination of blaMBLs.

Establishment of epidemiological cut-off values for cefoselis, a new fourth-generation cephalosporin, against Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, Proteus mirabilis and Pseudomonas aeruginosa.

OBJECTIVES: To establish the epidemiological cut-off values (ECOFFs) for cefoselis against Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae, Proteus mirabilis and Pseudomonas aeruginosa. METHODS: We collected 2288 non-repetitive clinical isolates from five laboratories throughout four cities in China. The cefoselis MICs and inhibition zone diameters for all isolates were established using the broth microdilution method and the disc diffusion method following EUCAST guidelines. MIC ECOFFs were determined by visual estimation and ECOFFinder software. Zone diameter ECOFFs were set if a high correlation of MICs and inhibition zone diameters was found by Pearson correlation. Zone diameter ECOFFs were finally determined by the visual estimate method. RESULTS: MICs of cefoselis were distributed from 0.008 to >256 mg/L for the four Enterobacterales species and from 0.25 to >256 mg/L for P. aeruginosa. MIC ECOFFs were 0.125 mg/L for E. coli, K. pneumoniae and P. mirabilis, 0.25 mg/L for E. cloacae and 32 mg/L for P. aeruginosa. A high correlation of MICs and zone diameters was observed for all Enterobacterales (|r| > 0.8, P < 0.001) and a relatively high correlation was found for P. aeruginosa (|r| = 0.71, P < 0.001). The zone diameter ECOFF was 24 mm for E. cloacae, E. coli and K. pneumoniae, 26 mm for P. mirabilis and 21 mm for P. aeruginosa. CONCLUSIONS: We determined MIC and zone diameter ECOFFs for cefoselis against four Enterobacterales species and P. aeruginosa. The establishment of ECOFFs for cefoselis provides clinicians with helpful guidance to differentiate WT and non-WT pathogens.