Project description:Transcriptomic analysis of Aspergillus fumigatus and Klebsiella pneumoniae grown as single and mixed biofilms. Overall design: RNA Sequencing was performed of Aspergillus fumigatus and Klebsiella pneumoniae grown as single and mixed biofilms for 24 hours at 37 °C in YPD medium. For mixed biofilms, Aspergillus fumigatus was pre-cultured for 12 hours to allow germination prior addition of Klebsiella pneumoniae. Cultures were harvested and three biological replicates were subjected for RNA isolation and sequencing library preparation.
Project description:Elucidating the RamA Regulon in Klebsiella pneumoniae and the transcriptome profiles of multidrug resistant Klebsiella pneumoniae Overall design: various strains of Klebsiella pneumoniae
Project description:Five Klebsiella pneumoniae isolates with reduced susceptibility to tigecycline (MIC, 2 microg/ml) were analyzed. A gene homologous to ramR of Salmonella enterica was identified in Klebsiella pneumoniae. Sequencing of ramR in the nonsusceptible Klebsiella strains revealed deletions, insertions, and point mutations. Transformation of mutants with wild-type ramR genes, but not with mutant ramR genes, restored susceptibility to tigecycline and repressed overexpression of ramA and acrB. Thus, this study reveals a molecular mechanism for tigecycline resistance in Klebsiella pneumoniae.
Project description:We describe here the genome sequence of the novel temperate Klebsiella pneumoniae phage KPP5665-2 isolated from a Klebsiella pneumoniae strain recovered from milk in Germany in 2016. The phage exhibited a narrow host range and a siphoviridal morphology. KPP5665-2-related prophage sequences were detected in whole-genome sequencing (WGS) data of various Klebsiella species isolates.
Project description:Klebsiella pneumoniae is a major threat to public health, causing significant morbidity and mortality worldwide. The emergence of highly drug-resistant strains is particularly concerning. There has been a recognition and division of Klebsiella pneumoniae into three distinct phylogenetic groups: Klebsiella pneumoniae, Klebsiella variicola, and Klebsiella quasipneumoniae. K. variicola and K. quasipneumoniae have often been described as opportunistic pathogens that have less virulence in humans than K. pneumoniae does. We recently sequenced the genomes of 1,777 extended-spectrum-beta-lactamase (ESBL)-producing K. pneumoniae isolates recovered from human infections and discovered that 28 strains were phylogenetically related to K. variicola and K. quasipneumoniae. Whole-genome sequencing of 95 additional non-ESBL-producing K. pneumoniae isolates recovered from patients found 12 K. quasipneumoniae strains. Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis initially identified all patient isolates as K. pneumoniae, suggesting a potential pitfall in conventional clinical microbiology laboratory identification methods. Whole-genome sequence analysis revealed extensive sharing of core gene content and plasmid replicons among the Klebsiella species. For the first time, strains of both K. variicola and K. quasipneumoniae were found to carry the Klebsiella pneumoniae carbapenemase (KPC) gene, while another K. variicola strain was found to carry the New Delhi metallo-beta-lactamase 1 (NDM-1) gene. K. variicola and K. quasipneumoniae infections were not less virulent than K. pneumoniae infections, as assessed by in-hospital mortality and infection type. We also discovered evidence of homologous recombination in one K. variicola strain, as well as one strain from a novel Klebsiella species, which challenge the current understanding of interrelationships between clades of Klebsiella. IMPORTANCEKlebsiella pneumoniae is a serious human pathogen associated with resistance to multiple antibiotics and high mortality. K. variicola and K. quasipneumoniae are closely related organisms that are generally considered to be less-virulent opportunistic pathogens. We used a large, comprehensive, population-based strain collection and whole-genome sequencing to investigate infections caused by these organisms in our hospital system. We discovered that K. variicola and K. quasipneumoniae isolates are often misidentified as K. pneumoniae by routine clinical microbiology diagnostics and frequently cause severe life-threatening infections similar to K. pneumoniae. The presence of KPC in K. variicola and K. quasipneumoniae strains as well as NDM-1 metallo-beta-lactamase in one K. variicola strain is particularly concerning because these genes confer resistance to many different beta-lactam antibiotics. The sharing of plasmids, as well as evidence of homologous recombination, between these three species of Klebsiella is cause for additional concern.
Project description:<i>Klebsiella</i> is a genus of Gram-negative bacteria known to be opportunistic pathogens that may cause a variety of infections in humans. Highly drug-resistant <i>Klebsiella</i> species, especially <i>K. pneumoniae</i>, have emerged rapidly and are becoming a major concern in clinical management. Although <i>K. pneumoniae</i> is considered the most important pathogen within the genus, the true clinical significance of the other species is likely underrecognized due to the inability of conventional microbiological methods to distinguish between the species leading to high rates of misidentification. Bacterial whole-genome sequencing (WGS) enables precise species identification and characterization that other technologies do not allow. Herein, we have characterized the diversity and traits of <i>Klebsiella</i> spp. in community-onset infections by WGS of clinical isolates (<i>n</i> = 105) collected during a prospective sepsis study in Sweden. The sequencing revealed that 32 of the 82 isolates (39.0%) initially identified as <i>K. pneumoniae</i> with routine microbiological methods based on cultures followed by matrix-assisted laser desorption-time of flight mass spectrometry (MALDI-TOF MS) had been misidentified. Of these, 23 were identified as <i>Klebsiella variicola</i> and nine as other members of the <i>K. pneumoniae</i> complex. Comparisons of the number of resistance genes showed that significantly fewer resistance genes were detected in <i>Klebsiella oxytoca</i> compared to <i>K. pneumoniae</i> and <i>K. variicola</i> (both values of <i>p</i> < 0.001). Moreover, a high proportion of the isolates within the <i>K. pneumoniae</i> complex were predicted to be genotypically multidrug-resistant (MDR; 79/84, 94.0%) in contrast to <i>K. oxytoca</i> (3/16, 18.8%) and <i>Klebsiella michiganensis</i> (0/4, 0.0%). All isolates predicted as genotypically MDR were found to harbor the combination of β-lactam, fosfomycin, and quinolone resistance markers. Multi-locus sequence typing (MLST) revealed a high diversity of sequence types among the <i>Klebsiella</i> spp. with ST14 (10.0%) and ST5429 (10.0%) as the most prevalent ones for <i>K. pneumoniae</i>, ST146 for <i>K. variicola</i> (12.0%), and ST176 for <i>K. oxytoca</i> (25.0%). In conclusion, the results from this study highlight the importance of using high-resolution genotypic methods for identification and characterization of clinical <i>Klebsiella</i> spp. isolates. Our findings indicate that infections caused by other members of the <i>K. pneumoniae</i> complex than <i>K. pneumoniae</i> are a more common clinical problem than previously described, mainly due to high rates of misidentifications.
Project description:Total RNA isolated from mid log grown cultures of K.pneumoniae and mutant strain in three independent times.Expression profile of K.pneumoniae and its pk muatant was compared. Overall design: Agilent one-color experiment,Organism: Klebsiella pneumoniae ,Agilent Custom Klebsiella pneumoniae 8x15k Microarray designed by Genotypic Technology Private Limited (AMADID: 079362)
Project description:Total RNA isolated from mid log grown cultures of K.pneumoniae and mutant strain in three independent times.Expression profile of K.pneumoniae and its RND muatant was compared. Overall design: Agilent one-color experiment,Organism: Klebsiella pneumoniae ,Agilent Custom Klebsiella pneumoniae 8x15k Microarray designed by Genotypic Technology Private Limited (AMADID: 079362)
Project description:Pathogens of the genus Klebsiella have been classified into distinct capsular (K) types for nearly a century. K typing of Klebsiella species still has important applications in epidemiology and clinical microbiology, but the serological method has strong practical limitations. Our objective was to evaluate the sequencing of wzi, a gene conserved in all capsular types of Klebsiella pneumoniae that codes for an outer membrane protein involved in capsule attachment to the cell surface, as a simple and rapid method for the prediction of K type. The sequencing of a 447-nucleotide region of wzi distinguished the K-type reference strains with only nine exceptions. A reference wzi sequence database was created by the inclusion of multiple strains representing K types associated with high virulence and multidrug resistance. A collection of 119 prospective clinical isolates of K. pneumoniae were then analyzed in parallel by wzi sequencing and classical K typing. Whereas K typing achieved typeability for 81% and discrimination for 94.4% of the isolates, these figures were 98.1% and 98.3%, respectively, for wzi sequencing. The prediction of K type once the wzi allele was known was 94%. wzi sequencing is a rapid and simple method for the determination of the K types of most K. pneumoniae clinical isolates.
Project description:A unique Klebsiella species strain, 10982, was cultured from a perianal swab specimen obtained from a patient in the University of Maryland Medical Center intensive care unit. Klebsiella sp. 10982 possesses a large IncA/C multidrug resistance plasmid encoding a novel FOX AmpC ?-lactamase designated FOX-10. A novel variant of the LEN ?-lactamase was also identified. Genome sequencing and bioinformatic analysis demonstrated that this isolate contains genes associated with nitrogen fixation, allantoin metabolism, and citrate fermentation. These three gene regions are typically present in either Klebsiella pneumoniae clinical isolates or Klebsiella nitrogen-fixing endophytes but usually not in the same organism. Phylogenomic analysis of Klebsiella sp. 10982 and sequenced Klebsiella genomes demonstrated that Klebsiella sp. 10982 is present on a branch that is located intermediate between the genomes of nitrogen-fixing endophytes and K. pneumoniae clinical isolates. Metabolic features identified in the genome of Klebsiella sp. 10982 distinguish this isolate from other Klebsiella clinical isolates. These features include the nitrogen fixation (nif) gene cluster, which is typically present in endophytic Klebsiella isolates and is absent from Klebsiella clinical isolates. Additionally, the Klebsiella sp. 10982 genome contains genes associated with allantoin metabolism, which have been detected primarily in K. pneumoniae isolates from liver abscesses. Comparative genomic analysis of Klebsiella sp. 10982 demonstrated that this organism has acquired genes conferring new metabolic strategies and novel antibiotic resistance alleles, both of which may enhance its ability to colonize the human body.