The emergence of carbapenemase-producing Citrobacter freundii poses a significant threat to public health worldwide. Here, we reported a C. freundii strain CWH001 which was resistant to all tested antimicrobials except tetracycline. Whole genome sequencing and analysis were performed. The strain, which belonged to a new sequence type ST139, showed close relationship with other foreign C. freundii strains through phylogenetic analysis. A novel variant of the intrinsic blaCMY gene located on the chromosome was identified and designated as blaCMY-152. Coexistence of blaNDM-1 with qnrS1 was found on a conjugative IncN plasmid, which had a backbone appearing in various plasmids. Other class A ESBL genes (blaVEB-3 and blaTEM-1) were also detected on two different novel plasmids. The emergence of multidrug-resistant C. freundii is of major concern, causing great challenges to the treatment of clinical infections. Great efforts need to be taken for the specific surveillance of this opportunistic pathogen.

Background Chlamydia psittaci is an avian pathogen that can cause lethal human infections. Diagnosis of C. psittaci pneumonia is often delayed due to nonspecific clinical presentations and limited laboratory diagnostic techniques. Results The MinION platform established the diagnosis in the shortest time, while BGISEQ-500 generated additional in-depth sequence data that included the rapid characterization of antibiotic susceptibility. Cytopathy appeared only in cell cultures of BALF. BALF yielded a higher bacterial load than sputum or blood, and may be the most suitable clinical specimen for the genomic diagnosis of severe pneumonia. Conclusions This study indicated that the benefits of metagenomic sequencing include rapid etiologic diagnosis of unknown infections and the provision of additional relevant information regarding antibiotic susceptibility. The continued optimization and standardization of sampling and metagenomic analysis promise to enhance the clinical utility of genomic diagnosis.

Rapid and accurate etiologic diagnosis accelerates targeted antimicrobial therapy. Metagenomic analysis has played a critical role in pathogen identification. In this study, we leveraged the advantages of both the MinION and BGISEQ-500 platforms to make a bacteriologic diagnosis from a culture-negative lung tissue sample from an immunocompromised patient with severe pneumonia. Real-time nanopore sequencing rapidly identified Klebsiella pneumoniae by an 823 bp specific sequence within 1 min. Genomic analysis further identified bla SHV-12 , bla KPC-2 , bla TEM-1 , bla CTX-M-65 , and other resistance genes. The same sample was further sequenced on the BGISEQ-500 platform, which presented consistent results regarding the most top dominant pathogens and provided additional information of resistance genes. Revised antibiotic treatment was followed by the patient's clinical recovery. Though sample preparation and the interpretation of final results still need to be improved further, metagenomic sequencing contributes to the accurate diagnosis of culture-negative infections and facilitates the rational antibiotic therapy.

The emergence of carbapenem-resistant Enterobacteriaceae poses a significant threat to public health worldwide. Here, we reported a multidrug-resistant Escherichia coli strain with two different bla NDM-5-carrying plasmids from China. Illumina short-read and MinION long-read whole genome sequencing were performed. Genomic analysis found that one bla NDM-5 gene together with mphA was located on a 55-kb IncX3 plasmid, while the other bla NDM-5 gene was on a novel 68-kb IncFII plasmid. Susceptibility testing and quantitative reverse transcription PCR results further indicated that the transconjugants with the IncX3 plasmid exhibited higher-level carbapenem resistance and expression of bla NDM-5 than those with both plasmids or the IncFII plasmid. Two other β-lactamase genes (bla CTX−M-15 and bla OXA-1) were also detected on another 160-kb IncF plasmid. This is the first report of coexistence of two bla NDM-5-carrying plasmids in a single bacterial isolate, highlighting the genetic complexity of NDM-5 carbapenemase circulation, and the urgent need for continued active surveillance.