Hao Xu

Objective: Carbapenem-resistant Klebsiella pneumoniae harboring bla _NDM poses a serious threat to public health; however, bla _NDM-15 remains poorly characterized outside the dominant epidemic lineages. Methods: We characterized K. pneumoniae strain ETFK6090, isolated from a perianal surveillance swab of an 11-month-old immunocompromised child in a paediatric intensive care unit. Investigations included antimicrobial susceptibility testing, broth conjugation, S1 nuclease PFGE with Southern blotting, complete genome sequencing, and comparative genomic analysis against 465 curated bla _NDM -positive K. pneumoniae genomes from 37 countries. Results: ETFK6090 belonged to ST580 and exhibited resistance to carbapenems, ceftazidime-avibactam, broad-spectrum cephalosporins, fluoroquinolones, gentamicin, chloramphenicol and trimethoprim-sulfamethoxazole; amikacin and fosfomycin retained low MICs. The complete genome comprised one chromosome and five plasmids, bla _NDM-15 was localized on a 46,161-bp IncX3 plasmid, confirmed by Southern blotting. Conjugation into Escherichia coli EC600 transferred carbapenem and cephalosporin resistance, confirming in vitro mobility. The bla _NDM-15 genetic environment retained a conserved bla _NDM module, with IS-mediated rearrangements at the downstream boundary. In the global comparison, bla _NDM-1 and bla _NDM-5 predominated, the ST580- bla _NDM-15 combination was exceedingly rare, and ETFK6090 constituted a distinct branch apart from major epidemic lineages. Conclusions: A transferable IncX3- bla _NDM-15 plasmid can emerge in an uncommon ST580 background, underscoring the necessity to extend genomic surveillance of carbapenem-resistant K. pneumoniae beyond dominant epidemic clones, particularly in high-risk paediatric and intensive-care settings.

Omadacycline and eravacycline are gradually being used as new tetracycline antibiotics for the clinical treatment of Gram-negative pathogens. Affected by various tetracycline-inactivating enzymes, there have been reports of resistance to eravacycline and omadacycline in recent years. We isolated a strain carrying the mobile tigecycline resistance gene tet (X4) from the feces of a patient in Zhejiang Province, China. The strain belongs to the rare ST485 sequence type. The isolate was identified as Klebsiella pneumoniae by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). The MICs of antimicrobial agents were determined using either the agar dilution method or the micro broth dilution method. The result showed that the isolate was resistant to eravacycline (MIC = 32 mg/L), omadacycline (MIC > 64 mg/L), and tigecycline (MIC > 32 mg/L). Whole-genome sequencing revealed that the tet (X4) resistance gene is located on the IncFII(pCRY) conjugative plasmid. tet (X4) is flanked by IS Vsa3 , and we hypothesize that this association contributes to the spread of the resistance gene. Plasmids were analyzed by S1-nuclease pulsed-field gel electrophoresis (S1-PFGE), Southern blotting, and electrotransformation experiment. We successfully transferred the plasmid carrying tet (X4) to the recipient bacteria by electrotransformation experiment. Compared with the DH-5α, the MICs of the transformant L3995-DH5α were increased by eight-fold for eravacycline and two-fold higher for omadacycline. Overall, the emergence of plasmid-borne tet (X4) resistance gene in a clinical isolate of K. pneumoniae ST485 underscores the essential requirement for the ongoing monitoring of tet (X4) to prevent and control its further dissemination in China. IMPORTANCE: There are still limited reports on Klebsiella pneumoniae strains harboring tetracycline-resistant genes in China, and K. pneumoniae L3995hy adds a new example to those positive for the tet (X4) gene. Importantly, our study raises concerns that plasmid-mediated resistance to omadacycline and eravacycline may spread further to a variety of ecological and clinical pathogens, limiting the choice of medication for extensively drug-resistant bacterial infections. Therefore, it is important to continue to monitor the prevalence and spread of tet (X4) and other tetracyclines resistance genes in K. pneumoniae and diverse bacterial populations.

The emerging infectious agent Elizabethkingia meningoseptica is associated with life-threatening infections in immunocompromised individuals. However, there are limited data on its geographic distribution, phylogenetic evolution, pathogenesis, and transmission. In this study, we comprehensively analyze and compare the genomic features, evolutionary history, emergence date, and transmission networks of global E. meningoseptica . Geographical distribution reveals the presence of the emerging bacteria in Asia, Europe, and North America, three continents with similar latitudes. Phylogenetic analyses show no relationship between the strain’s evolutionary history and its location, origin, or source, despite the presence of genetic diversity. Analysis of the emergence timeline suggests that America is the most likely source of E. meningoseptica with the common ancestor of this pathogen dating back 90 years. Putative transmission networks indicate that E. meningoseptica bacteria can spread within the same hospital and even across borders. Minor variations in resistance genotypes and virulence genes are observed, supporting existing evidence of inherent resistance and pathogenicity in E. meningoseptica . Additionally, minocycline and doxycycline demonstrate potent antimicrobial activity against this pathogen, making them promising candidates for treating E. meningoseptica infections. Our research highlights the potential for severe nosocomial outbreaks caused by E. meningoseptica with horizontal transmission occurring between countries worldwide. To prevent future outbreak infections, increased genomic surveillance of global E. meningoseptica populations is necessary. IMPORTANCE: Elizabethkingia meningoseptica is an emerging infectious agent associated with life-threatening infections in immunocompromised individuals. However, there are limited data available on the genomic features of E. meningoseptica . This study aims to characterize the geographical distribution, phylogenetic evolution, pathogenesis, and transmission of this bacterium. A systematic analysis of the E. meningoseptica genome revealed that a common ancestor of this bacterium existed 90 years ago. The evolutionary history showed no significant relationship with the sample source, origin, or region, despite the presence of genetic diversity. Whole genome sequencing data also demonstrated that E. meningoseptica bacteria possess inherent resistance and pathogenicity, enabling them to spread within the same hospital and even across borders. This study highlights the potential for E. meningoseptica to cause severe nosocomial outbreaks and horizontal transmission between countries worldwide. The available evidence is crucial for the development of evidence-based public health policies to prevent global outbreaks caused by emerging pathogens.

Background: Late-onset group B Streptococcus (LOGBS) sepsis is a cause of infection and death in infants. Infected breast milk has been considered a source of neonatal GBS infection and invasive infection. However, mother-to-infant transmission of GBS detected by the high-resolution diagnostic method is rarely reported. Methods: This study describes a low-weight premature infant who developed late-onset GBS septicemia 21 days after birth. GBS strains isolated from the mother’s cervical secretion, the mother’s milk, and the baby’s blood were cultured to identify the source of GBS infection. We further confirmed the GBS isolates through matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF-MS). Finally, we performed whole-genome sequencing (WGS) and phylogenetic analyses on the GBS strains recovered. Results: GBS isolates were cultured from the bloodstream of the premature infant and the mother’s milk, respectively. Subsequently, WGS and phylogenetic analyses on three GBS isolates demonstrated that the GBS strain from the infant’s bloodstream was 100% homologous to that from the mother’s breast milk, which had some different gene fragments from the GBS strain from the mother’s cervical secretion. It provided evidence that this infant’s late-onset GBS septicemia originated from his mother’s breast milk instead of the vertical mother-to-infant transmission. Conclusion: Through WGS and phylogenetic analysis of the GBS strains, we proved in this study that the late-onset GBS sepsis in a premature infant was derived from his mother’s breast milk. It indicated that WGS diagnosis is an effective tool for infection tracing. Furthermore, this report provides direction for preventing late-onset GBS infection.

Purpose: To investigate the phenotypic and genomic characteristics of the multi-drug resistant and hypervirulent Klebsiella pneumoniae strain recovered from bacteremia. Methods: Antimicrobial susceptibility testing (AST) was performed by the microdilution method. Antimicrobial resistance genes, virulence-associated genes, multilocus sequence typing (MLST), and plasmid replicon were characterized by next-generation sequencing (NGS) and nanopore sequencing. S1 nuclease-pulsed field gel electrophoresis (S1-PFGE) and Southern blotting were performed to characterize the plasmid profile. Results: The hypervirulent colistin- and carbapenem-resistant K. pneumoniae strain DY2009 was identified as ST5571, co-carrying mcr-1, bla_NDM-1, and bla_OXA-10. In silico analysis found that it was K2 serotype. AST results revealed that DY2009 was resistant to carbapenems, cephalosporins, ciprofloxacin, chloramphenicol, and colistin but remained susceptible to aztreonam, gentamicin, amikacin, and tigecycline. Through the whole-genome analysis, a variety of virulence determinants were identified, including rmpA. Plasmid analysis confirmed that the mcr-1 and bla_NDM-1 gene harbored a ∼ 33 kb IncX4 plasmid and a ∼ 44 kb IncX3 plasmid. In contrast, bla_OXA-10 was encoded by chromosome. Conclusion: To the best of our knowledge, we first report the clinical hypervirulent K. pneumoniae isolate co-producing MCR-1, NDM-1, and OXA-10 causing bacteremia. We found that mcr-1 and bla_NDM-1 genes were located on two self-conjugative epidemic plasmids, contributing to the widespread MCR-1 and NDM-1 in China. The results of this work improve our understanding of the genetic background of colistin- and carbapenem-resistant K. pneumoniae isolate from bacteremia and the resistance mechanisms. Our findings highlight the urgent need for infection control of such strain to prevent it from becoming an extensive-drug resistant clone.

Background: To investigate the antimicrobial profiles and genomic characteristics of MDR-Citrobacter spp. strains isolated from Fennec fox imported from Sudan to China. Methods: Four Citrobacter spp. strains were isolated from stool samples. Individual fresh stool samples were collected and subsequently diluted in phosphate buffered saline as described previously. The diluted fecal samples were plated on MacConkey agar supplemented with 1 mg/l cefotaxime and incubated for 20 h at 37 °C. Matrix-assisted laser desorption/ionization–time of flight mass spectrometry (MALDI–TOF–MS) was used for identification. Antimicrobial susceptibility testing was performed using the broth microdilution method. Whole-genome sequencing was performed on an Illumina Novaseq-6000 platform. Acquired antimicrobial resistance genes and plasmid replicons were detected using ResFinder 4.1 and PlasmidFinder 1.3, respectively. Comparative genomic analysis of 277 Citrobacter genomes was also performed. Results: Isolate FF141 was identified as Citrobacter cronae while isolate FF371, isolate FF414, and isolate FF423 were identified as Citrobacter braakii. Of these, three C. braakii isolates were further confirmed to be extended-spectrum β-lactamases (ESBL)-producer. All isolates are all multidrug resistance (MDR) with resistance to multiple antimicrobials. Plasmid of pKPC-CAV1321 belong to incompatibility (Inc) group. Comparative genomics analysis of Citrobacter isolates generated a large core-genome. Genetic diversity was observed in our bacterial collection, which clustered into five main clades. Human, environmental and animal Citrobacter isolates were distributed into five clusters. Conclusions: To our knowledge, this is the first investigation of MDR-Citrobacter from Fennec Fox. Our phenotypic and genomic data further underscore the threat of increased ESBL prevalence in wildlife and emphasize that increased effort should be committed to monitoring the potentially rapid dissemination of ESBL-producers with one health perspective.

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