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Objectives Routine blood examination, one of the most commonly performed tests in clinical laboratories, directly reflects the overall state of the body such as inflammation, anemia, and thrombocytopenia. The accuracy of these indicators by tests may be perturbed by various factors including anticoagulants, antibodies, and temperatures. Pseudoleukopenia caused by leukoagglutination was rarely described in the literature. Case presentation We report a rare and unusual pseudo-leukopenia case of a 75-year-old female with a stroke. Blood samples from the patient were collected using different anticoagulants and determined the hematologic parameters and blood smears. We observed the extent of leukocyte aggregation at different anticoagulants or temperatures. The intensity of leukoagglutination was attenuated after incubating at 37 °C for 30 min. After anti-infection treatment and symptomatic treatment, the leukoagglutination of the patient gradually weakened. Conclusions We have found the reason for the pseudo-leukopenia and the leukocyte aggregation phenomenon may vary with disease progression.

Carbapenem-resistant and hypervirulent K. pneumoniae (CR-HvKP) strains that have emerged recently have caused infections of extremely high mortality in various countries. In this study, we discovered a conjugative plasmid that encodes carbapenem resistance and hypervirulence in a clinical ST86 K2 CR-HvKP, namely 17ZR-91. The conjugative plasmid (p17ZR-91-Vir-KPC) was formed by fusion of a non-conjugative pLVPK-like plasmid and a conjugative blaKPC-2-bearing plasmid and is present dynamically with two other non-fusion plasmids. Conjugation of p17ZR-91-Vir-KPC to other K. pneumoniae enabled them to rapidly express the carbapenem resistance and hypervirulence phenotypes. More importantly, genome analysis provided direct evidence that p17ZR-91-Vir-KPC could be directly transmitted from K2 CR-HvKP strain, 17ZR-91, to ST11 clinical K. pneumoniae strains to convert them into ST11 CR-HvKP strains, which explains the evolutionary mechanisms of recently emerged ST11 CR-HvKP strains., Carbapenem-resistant and hypervirulent Klebsiella pneumoniae strains are emerging. Here Xie et al. show that these phenotypes are carried on a plasmid formed from the fusion of a virulence plasmid with a conjugative plasmid.

Objectives To investigate the genomic and phenotypic characteristics of an MDR Empedobacter falsenii strain isolated from a Chinese patient, which was phenotypically resistant to all last-line antibiotics (carbapenems, colistin and tigecycline). Methods Species identity was determined by MALDI-TOF MS analysis. The complete genome sequence of the isolate was determined by WGS and the genetic elements conferring antimicrobial resistance were determined. The origin of this strain was tracked by phylogenetic analysis. Results The E. falsenii strain was genetically most closely related to an Empedobacter sp. strain isolated from the USA. Members of E. falsenii are speculated to be intrinsically resistant to colistin. The carbapenem resistance of this strain was conferred by a chromosomal blaEBR-2 variant gene. Phylogenetic analysis indicated that the gene encoding the EBR β-lactamase was widely distributed in Empedobacter spp. Tigecycline resistance was mediated by a tet(X) variant gene encoded by a non-conjugative and non-typeable plasmid. Conclusions The MDR phenotype of the E. falsenii isolate was conferred by different mechanisms. Findings from us and others indicate that E. falsenii may serve as a reservoir for carbapenem and tigecycline resistance determinants.

Background: Carbapenem-resistant K. pneumoniae (CRKP) is the leading cause of hospital infections. The prevalence of CRKP in human and the risk factors that predispose occurrence of CRKP infections need to be determined. Methods: We conducted a retrospective, cross-sectional study of K. pneumoniae strains collected in a hospital in Zhejiang Province, China, from Jan 2014 to Dec 2017. The strains were characterized by antimicrobial susceptibility tests and whole genome sequencing. Then we investigated risk factors of CRKP infections and depicted the likely disease outcomes. Findings: 804 patients tested positive for intestinal CRKP or carbapenem susceptible strains (CSKP) in total. Ninety-seven patients were positive for both intestinal and extra-intestinal CRKP and had significantly (P

Plasmid-mediated, transmissible, tigecycline-inactivating enzyme Tet(X) has attracted considerable public attention. However, so far studies have not addressed its impact on public health and the ecosystem. Herein, we report the prevalence and molecular epidemiology of tet(X)-positive bacteria (TPB) from diverse sources, investigate the host-specificity of TPB and the transferability of tet(X). Sample collection was conducted between 2018 and 2020 in 30 provinces in China. PCR screening suggested tet(X) was prevalent among freshwater fishes (24.7%, 95% CI 19.4–30.7%), followed by chickens (23.6%, 21.2–26.2%), cattle (19.3%, 16.4–22.5%), healthy individuals (6.2%, 5.4–7.1%), and patients (0.3%, 0.0–1.1%). Soil and freshwater samples all tested negative for tet(X). A total of 289 TPB were isolated from 7516 samples (120/1181 chicken, 82/669 cattle, 68/3229 healthy individual, 17/239 freshwater fish and 2/2121 clinical samples). TPB distributed in six major families of bacteria including Moraxellaceae (n = 99, 34.3%), Flavobacteriaceae (n = 95, 32.9%), Enterobacteriaceae (n = 83, 28.7%), Pseudomonadaceae (n = 9, 3.1%), Sphingobacteriaceae (n = 2, 0.7%) and unclassified Gammaproteobacteria (n = 1, 0.3%). Diverse tet(X) genes including tet(X2), tet(X3), tet(X4), tet(X5) and tet(X6) were identified from different TPB. The tet(X)-positive bacteria were highly diverse, with ST10 complex belonging to the dominant E. coli clone. Novel hosts of tet(X) including Enterobacter hormaechei, Ignatzschineria indica and Oblitimonas alkaliphila were identified. Isolates from different families exhibited different antimicrobial resistance profiles. Co-existence of tet(X) with other resistance genes such as floR (66.8%) and carbapenemase genes (33.2%) was commonly observed. tet(X) could be transferred among E. coli isolates at frequencies from 10−4 to 10−10. Species other than E. coli failed to transfer tet(X) gene to the E. coli recipient via conjugation. Discriminant analysis of principal components analysis suggested inter-host transmission of tet(X)-positive E. coli among diverse hosts was not observed. Future studies are needed to monitor the transmission trend as well as the impact of this resistance gene in clinical infection control.

Summary Background Hypervirulent Klebsiella pneumoniae strains often cause life-threatening community-acquired infections in young and healthy hosts, but are usually sensitive to antibiotics. In this study, we investigated a fatal outbreak of ventilator-associated pneumonia caused by a new emerging hypervirulent K pneumoniae strain. Methods The outbreak occurred in the integrated intensive care unit of a new branch of the Second Affiliated Hospital of Zhejiang University (Hangzhou, China). We collected 21 carbapenem-resistant K pneumoniae strains from five patients and characterised these strains for their antimicrobial susceptibility, multilocus sequence types, and genetic relatedness using VITEK-2 compact system, multilocus sequence typing, and whole genome sequencing. We selected one representative isolate from each patient to establish the virulence potential using a human neutrophil assay and Galleria mellonella model and to establish the genetic basis of their hypervirulence phenotype. Findings All five patients had undergone surgery for multiple trauma and subsequently received mechanical ventilation. The patients were aged 53–73 years and were admitted to the intensive care unit between late February and April, 2016. They all had severe pneumonia, carbapenem-resistant K pneumoniae infections, and poor responses to antibiotic treatment and died due to severe lung infection, multiorgan failure, or septic shock. All five representative carbapenem-resistant K pneumoniae strains belonged to the ST11 type, which is the most prevalent carbapenem-resistant K pneumoniae type in China, and originated from the same clone. The strains were positive on the string test, had survival of about 80% after 1 h incubation in human neutrophils, and killed 100% of wax moth larvae ( G mellonella ) inoculated with 1 × 10 6 colony-forming units of the specimens within 24 h, suggesting that they were hypervirulent K pneumoniae . Genomic analyses showed that the emergence of these ST11 carbapenem-resistant hypervirulent K pneumoniae strains was due to the acquisition of a roughly 170 kbp pLVPK-like virulence plasmid by classic ST11 carbapenem-resistant K pneumoniae strains. We also detected these strains in specimens collected in other regions of China. Interpretation The ST11 carbapenem-resistant hypervirulent K pneumoniae strains pose a substantial threat to human health because they are simultaneously hypervirulent, multidrug resistant, and highly transmissible. Control measures should be implemented to prevent further dissemination of such organisms in the hospital setting and the community. Funding Chinese National Key Basic Research and Development Program and Collaborative Research Fund of Hong Kong Research Grant Council.

We report the identification of a carbapenem-resistant, hypervirulent Klebsiella pneumoniae (hvKp) strain which produced the carbapenemase VIM-1. Genomic analysis showed that the strain belonged to sequence type ST23 and serotype K1, a major hvKp clone, and harbored three resistance-encoding plasmids. Among them, a bla VIM-1 -bearing plasmid was found to possess a mosaic structure presumably generated by multiple gene mobilization events.

This study reported the clonal dissemination of OXA-232-producing sequence type 15 (ST15) carbapenem-resistant Klebsiella pneumoniae among elderly patients in China. All patients were immunocompromised, suffered from multiple underlying diseases, and were hospitalized for a prolonged period; however, they slowly recovered on antimicrobial therapy. The bla(OXA-232) gene was in a 6.1-kb ColKP3-type nonconjugative plasmid. The strains displayed a multidrug resistance phenotype and were not hypervirulent despite harboring a virulence plasmid. Active surveillance should be enforced to control further transmission.

Gastrointestinal carriage is regarded as a major reservoir of K. pneumoniae infections, especially in intensive care patients. A total of 101 (95.3%) KPC-producing carbapenem-resistant K. pneumoniae (CRKP) isolates were identified among 106 CRKP isolates collected from stool samples of inpatients performing active rectal screening for carbapenem-resistant Enterobacteriaceae during hospitalization in the ICUs of a tertiary hospital between 2016 and 2017. Among them, six KPC-producing CRKP isolates from three patients (two isolates for each patient) were identified with distinct antibacterial susceptibility. Our findings showed that: (1) blaKPC–2 gene is predominant in CRKP strains isolated from the intensive care patients and can be incorporated into various plasmids that are transmissible among multiple bacterial hosts in the human gastrointestinal tract; (2) the human gastrointestinal tract has a capacity to dynamically colonize multiple clones of CRKP strains with varied plasmids, diverse antimicrobial resistance genes and virulence genes. K. pneumoniae colonization is an important step in progression to extraintestinal infection, which provides the rationale for establishing intervention measures to prevent subsequent infection. Thus, close surveillance on CRKP colonization, together with effective infection prevention and control measures, should be put into practice.

Background\ud \ud Carriage of carbapenem-resistant Enterobacteriaceae (CRE) in humans may contribute to the dissemination of CRE and impact on communities and healthcare facilities. Carbapenem-resistant Escherichia coli (CREC) is one of the major type of CRE in the human gut. Here, we describe a cross-sectional study to investigate the prevalence of CREC, and in particular the mcr-1 carrying CREC, in health volunteers in China.\ud \ud \ud Methods\ud \ud During September to December 2016, 3859 non-duplicated stool specimens were collected from healthy volunteers who received regular physical examinees in healthcare centers located in 19 provinces across China. Enrichment culture supplemented meropenem was used to isolate CREC. Carbapenemase producing determinants and the mcr-1 gene were determined by PCR amplification and sequencing. Isolates were further analyzed by antibiotic susceptibility test, genotyping, and whole genome analysis.\ud \ud \ud Findings\ud \ud A total of 92 non-duplicated CREC were isolated from 3859 stool specimens, among which 43 CREC are carbapenemase positive. In addition, the co-existence of blaNDM and mcr-1 was found in 14 CREC, which also showed resistance to the majority of all antimicrobial agents analyzed. The genetic background of these CREC isolates are highly diversified based on molecular typing. Furthermore, whole genome sequence indicated that NDM-5 is the predominant determinant conferring carbapenem resistance in CREC, and that NDM-5 carrying plasmids (IncX3) are very similar.\ud \ud \ud Interpretation\ud \ud The incidence of CREC carriage in healthy people in China was small; however, the co-existence of CREC with mcr-1 is disconcerting. Therefore, pre-screening prior to admission and monitoring of patients on high-dependency wards is highly recommended to control and prevent the dissemination of CRE in hospitals.\ud \ud \ud Outstanding Question\ud \ud The high prevalence of CREC in the healthy people should not be underestimated, as it may increase the risk of infection. This knowledge could have impact on the pre-screening and monitoring of CRE before patient administration.

The global spread of carbapenem-resistant Enterobacteriaceae (CRE) is one of the most severe threats to human health in a clinical setting. The recent emergence of plasmid-mediated colistin resistance gene mcr-1 among CRE strains greatly compromises the use of colistin as a last resort for the treatment of infections caused by CRE. This study aimed to understand the current epidemiological trends and characteristics of CRE from a large hospital in Henan, the most populous province in China. From 2014 to 2016, a total of 7,249 Enterobacteriaceae isolates were collected from clinical samples, among which 18.1% (1,311/7,249) were carbapenem resistant. Carbapenem-resistant Klebsiella pneumoniae and carbapenem-resistant Escherichia coli were the two most common CRE species, with Klebsiella pneumoniae carbapenemases (KPC) and New Delhi metallo-β-lactamases (NDM), respectively, responsible for the carbapenem resistance of the two species. Notably, >57.0% ( n = 589) of the K. pneumoniae isolates from the intensive care unit were carbapenem resistant. Furthermore, bla NDM-5 and mcr-1 were found to coexist in one E. coli isolate, which exhibited resistance to almost all tested antibiotics. Overall, we observed a significant increase in the prevalence of CRE isolates during the study period and suggest that carbapenems may no longer be considered to be an effective treatment for infections caused by K. pneumoniae in the studied hospital.

The rapid dissemination of non-conjugative virulence plasmids among non-K1/K2 types of Klebsiella pneumoniae poses an unprecedented threat to human health, yet the underlying mechanisms governing dissemination of such plasmids is unclear. In this study, a novel 68 581 bp IncFIA plasmid is discovered that can be fused to a hypervirulence-encoding plasmid to form a hybrid conjugative virulence plasmid in conjugation experiments; such fusion events involve homologous recombination between a 241 bp homologous region located in each of the two plasmids. The fusion hypervirulence-encoding plasmid can be conjugated to both classic and blaKPC-2 -bearing carbapenem-resistant K. pneumoniae strains through conjugation, enabling such strains to acquire the ability to express the hypervirulence phenotype. Dissemination of this fusion virulence plasmid will impose an enormous burden on current efforts to control and treat infections caused by multidrug resistant and hypervirulent K. pneumoniae.