Spatial Variations in Growth Rate within Klebsiella pneumoniae Colonies and Biofilm

1996 ◽  
Vol 12 (3) ◽  
pp. 316-321 ◽  
Author(s):  
E.J. Wentland ◽  
P.S. Stewart ◽  
C.-T. Huang ◽  
G.A. McFeters
Keyword(s):  
Growth Rate ◽  
Klebsiella Pneumoniae ◽  
Spatial Variations

Glycerol kinase as a substitute for dihydroxyacetone kinase in a mutant of Klebsiella pneumoniae

1982 ◽  
Vol 152 (3) ◽  
pp. 1303-1307
Author(s):  
R Z Jin ◽  
R G Forage ◽  
E C Lin
Keyword(s):  
Energy Source ◽  
Growth Rate ◽  
Klebsiella Pneumoniae ◽  
Sole Source ◽  
Glycerol Kinase ◽  
Aerobic Growth ◽  
Dihydroxyacetone Kinase ◽  
Rate Limiting

With dihydroxyacetone as the sole source of carbon and energy, constitutively synthesized glycerol kinase of the glp system supported aerobic growth of Klebsiella pneumoniae mutants lacking the inducible dihydroxyacetone kinase of the dha system. Glycerol kinase had an apparent Km of 0.01 mM for its physiological substrate and 1 mM for its surrogate substrate. However, the growth rate on dihydroxyacetone of cells relying on glycerol kinase increased with the concentration of the carbon and energy source up to 50 mM, suggesting that permeation is rate limiting.

scholarly journals 494. Fitness Cost of mcr-1-Mediated Colistin Resistance in Carbapenemase-Producing Klebsiella pneumoniae

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S241-S241
Author(s):  
Yi-Tsung Lin ◽  
Yi-Hsiang Cheng ◽  
Sheng-Hua Chou ◽  
Ying-Chi Huang ◽  
Liang Chen
Keyword(s):  
Growth Rate ◽  
Klebsiella Pneumoniae ◽  
Whole Genome Sequencing ◽  
Genome Sequencing ◽  
Plasmid Stability ◽  
Fatal Case ◽  
Fitness Cost ◽  
Whole Genome ◽  
Colistin Resistance ◽  
The Impact

Abstract Background The emergence of mobile colistin resistance gene mcr-1, a plasmid-borne polymyxin resistance mechanism, in carbapenem-resistant Klebsiella pneumoniae is an alarming concern. However, previous studies showed that the acquisition of mcr-1 was associated with a significant biological fitness cost in K. pneumoniae. We aimed to study the impact of mcr-1 on the biological fitness in clinical carbapenemase-producing K. pneumoniae strains. Methods Clinical carbapenemase-producing K. pneumoniae strains were collected consecutively at the Taipei Veterans General Hospital between November 2017 and December 2018. The strain positive for mcr-1 was subjected to whole-genome sequencing to delineate its genomic features. Escherichia coli J53 strain was used as the recipient strain in plasmid conjugation assay and the transconjugants were selected with sodium azide and colistin. Plasmid stability was tested by serial passaging in antibiotic-free LB broth for 28 days. The growth rate was compared between the parental mcr-1-bearing strain and the plasmid-cured strain. Results One ST11 strain isolated from a fatal case with bacteremia (KP2509) was found to harbor blaKPC-2, blaOXA-48, and mcr-1. This strain was resistant to colistin (MIC=8 mg/L) and imipenem (MIC≥16 mg/L). Whole-genome sequencing of KP2509 showed that mcr-1, blaKPC-2 and blaOXA-48 were located on an IncHI-FIB type plasmid of 319 Kb, an IncFII type plasmid of 96 Kb, and an IncL type plasmid of 64 Kb, respectively. Conjugation efficiency of mcr-1-bearing plasmid was 2.24 × 10–4, and the colistin MIC of E. coli J53 transconjugant increased from 0.5 to 8 mg/L. The mcr-1-bearing plasmid in KP2509 showed high plasmid stability, and only ~1% were lost after 27-day passages. The resulting plasmid-cured strain (PC-KP2509) was susceptible to colistin (MIC=0.5 mg/L) and had a similar growth rate to that of parental mcr-1-bearing strain KP2509. Conclusion We identified an ST11 K. pneumoniae strain carrying blaKPC-2, blaOXA-48, and mcr-1 genes causing a fatal bacteremia. The large mcr-1-bearing plasmid confers a moderate level of colistin resistance but without significant biological fitness cost in carbapenemase-producing K. pneumoniae, which could result in a serious threat clinically. Disclosures All authors: No reported disclosures.

scholarly journals Role of Nutrient Limitation and Stationary-Phase Existence in Klebsiella pneumoniae Biofilm Resistance to Ampicillin and Ciprofloxacin

2003 ◽  
Vol 47 (4) ◽  
pp. 1251-1256 ◽  
Author(s):  
Jeff N. Anderl ◽  
Jeff Zahller ◽  
Frank Roe ◽  
Philip S. Stewart
Keyword(s):  
Growth Rate ◽  
Klebsiella Pneumoniae ◽  
Stationary Phase ◽  
Specific Growth Rate ◽  
Nutrient Limitation ◽  
Prolonged Exposure ◽  
Specific Growth ◽  
Nitrogen Sources ◽  
Biofilm Bacteria ◽  
Bacterial Aggregates

ABSTRACT Biofilms formed by Klebsiella pneumoniae resisted killing during prolonged exposure to ampicillin or ciprofloxacin even though these agents have been shown to penetrate bacterial aggregates. Bacteria dispersed from biofilms into medium quickly regained most of their susceptibility. Experiments with free-floating bacteria showed that stationary-phase bacteria were protected from killing by either antibiotic, especially when the test was performed in medium lacking carbon and nitrogen sources. These results suggested that the antibiotic tolerance of biofilm bacteria could be explained by nutrient limitation in the biofilm leading to stationary-phase existence of at least some of the cells in the biofilm. This mechanism was supported by experimental characterization of nutrient availability and growth status in biofilms. The average specific growth rate of bacteria in biofilms was only 0.032 h−1 compared to the specific growth rate of planktonic bacteria of 0.59 h−1 measured in the same medium. Glucose did not penetrate all the way through the biofilm, and oxygen was shown to penetrate only into the upper 100 μm. The specific catalase activity was elevated in biofilm bacteria to a level similar to that of stationary-phase planktonic cells. Transmission electron microscopy revealed that bacteria were affected by ampicillin near the periphery of the biofilm but were not affected in the interior. Taken together, these results indicate that K. pneumoniae in this system experience nutrient limitation locally within the biofilm, leading to zones in which the bacteria enter stationary phase and are growing slowly or not at all. In these inactive regions, bacteria are less susceptible to killing by antibiotics.

Osmoregulation in Klebsiella pneumoniae: enhancement of anaerobic growth and nitrogen fixation under stress by proline betaine, γ-butyrobetaine, and other related compounds

1984 ◽  
Vol 30 (3) ◽  
pp. 299-305 ◽  
Author(s):  
Daniel Le Rudulier ◽  
Theophile Bernard ◽  
Gabrielle Goas ◽  
Jack Hamelin
Keyword(s):  
Growth Rate ◽  
Nitrogen Fixation ◽  
Klebsiella Pneumoniae ◽  
Osmotic Stress ◽  
Anaerobic Growth ◽  
Methyl Groups ◽  
Wild Type ◽  
Whole Cells ◽  
Related Compounds ◽  
Exogenous Proline

Exogenous proline betaine (stachydrine or N-dimethylproline) or γ-butyrobetaine (γ-trimethylaminobutyrate), at a concentration as low as 1 mM, were found to stimulate the growth rate of Klebsiella pneumoniae, wild type M5A1, in media of inhibitory osmotic strength (0.8 M NaCl). Simultaneously, nitrogen fixation by whole cells, a process particularly sensitive to osmotic stress, was strongly enhanced by these compounds. However, in the absence of sodium chloride, both the growth and nitrogen fixation were not affected by the addition of the methylammonium derivatives in the medium. The sensitivity of the nitrogen fixation to osmotic stress was used as a bioassay to evaluate the potentiality of osmoprotective compound in relation to the number of methyl groups on the nitrogen atom of glycine, proline, and γ-aminobutyrate. Experiments with sarcosine (monomethylglycine), dimethylglycine, and glycine betaine (trimethylglycine), or experiments with mono- and di-methylproline or γ-mono-, γ-di-, γ-tri-methylaminobutyrate, indicated that the greatest stress tolerance was always obtained with the more N-methylated compounds.

scholarly journals Characterization of a mcr-1 and CRISPR-Cas System Co-harboring Plasmid in a Carbapenemase-Producing High-Risk ST11 Klebsiella pneumoniae Strain

2021 ◽  
Vol 12 ◽  
Author(s):  
Yi-Hsiang Cheng ◽  
Sheng-Hua Chou ◽  
Po-Han Huang ◽  
Tsuey-Ching Yang ◽  
Yu-Fan Juan ◽  
...  
Keyword(s):  
Growth Rate ◽  
Klebsiella Pneumoniae ◽  
Plasmid Stability ◽  
Medical Center ◽  
Conjugative Plasmid ◽  
Type Iv ◽  
Positive Strain ◽  
Stable Plasmid

We set out to study the prevalence of the mcr-1 gene in carbapenemase-producing Klebsiella pneumoniae (CPKP) strains, and to determine whether its presence is associated with a fitness cost. A total of 234 clinical CPKP isolates were collected from a tertiary medical center in Taiwan from January 2018 to January 2019. The mcr-1 and carbapenemase genes were detected by polymerase chain reaction (PCR) followed by Sanger sequencing. The mcr-1-positive carbapenemase-producing strain was characterized by whole genome sequencing, a plasmid stability test and a conjugation assay. In vitro growth rate and an in vivo virulence test were compared between the parental mcr-1-positive strain and its mcr-1 plasmid-cured strain. We identified only one mcr-1 positive strain (KP2509), co-harboring blaKPC–2 and blaOXA–48, among 234 (1/234, 0.43%) CPKP strains. KP2509 and its Escherichia coli mcr-1 transconjugant showed moderate colistin resistance (MIC = 8 mg/L). The mcr-1 is located on a large conjugative plasmid (317 kb), pKP2509-MCR, with three replicons, IncHI, IncFIB, and IncN. Interestingly, a complete Type IV-A3 CRISPR-Cas system was identified in pKP2509-MCR. Plasmid pKP2509-MCR was highly stable in KP2509 after 270 generation of passage, and the pKP2509-MCR cured strain PC-KP2509 showed similar growth rate and in vivo virulence in comparison to KP2509. The prevalence of mcr-1 in CPKP strains remains low in our center. Notably, we identified a large plasmid with multiple replicons containing both the mcr-1 and the Type IV-3A CRISPR-Cas genes. The further spread of this highly stable plasmid raises concern that it may promote the increase of mcr-1 prevalence in CPKP.

scholarly journals Expanded Role for the Nitrogen Assimilation Control Protein in the Response of Klebsiella pneumoniae to Nitrogen Stress

2010 ◽  
Vol 192 (19) ◽  
pp. 4812-4820 ◽  
Author(s):  
Ryan L. Frisch ◽  
Robert A. Bender
Keyword(s):  
Growth Rate ◽  
Stress Response ◽  
Klebsiella Pneumoniae ◽  
Nitrogen Metabolism ◽  
Nitrogen Limitation ◽  
Nitrogen Assimilation ◽  
Nitrogen Sources ◽  
Nitrogen Stress ◽  
A Genome ◽  
Regulated Promoters

ABSTRACT Klebsiella pneumoniae is able to utilize many nitrogen sources, and the utilization of some of these nitrogen sources is dependent on the nitrogen assimilation control (NAC) protein. Seven NAC-regulated promoters have been characterized in K. pneumoniae, and nine NAC-regulated promoters have been found by microarray analysis in Escherichia coli. So far, all characterized NAC-regulated promoters have been directly related to nitrogen metabolism. We have used a genome-wide analysis of NAC binding under nitrogen limitation to identify the regions of the chromosome associated with NAC in K. pneumoniae. We found NAC associated with 99 unique regions of the chromosome under nitrogen limitation. In vitro, 84 of the 99 regions associate strongly enough with purified NAC to produce a shifted band by electrophoretic mobility shift assay. Primer extension analysis of the mRNA from genes associated with 17 of the fragments demonstrated that at least one gene associated with each fragment was NAC regulated under nitrogen limitation. The large size of the NAC regulon in K. pneumoniae indicates that NAC plays a larger role in the nitrogen stress response than it does in E. coli. Although a majority of the genes with identifiable functions that associated with NAC under nitrogen limitation are involved in nitrogen metabolism, smaller subsets are associated with carbon and energy acquisition (18 genes), and growth rate control (10 genes). This suggests an expanded role for NAC regulation during the nitrogen stress response, where NAC not only regulates genes involved in nitrogen metabolism but also regulates genes involved in balancing carbon and nitrogen pools and growth rate.

scholarly journals The effect of pH on bacterial growth and histamine formation by Klebsiella pneumoniae CK02 and Raoultella ornithinolytica TN01

2021 ◽  
Vol 919 (1) ◽  
pp. 012039
Author(s):  
R Alya’ainun ◽  
E Y Fathoni ◽  
I D Puspita
Keyword(s):  
Growth Rate ◽  
Klebsiella Pneumoniae ◽  
Optimal Growth ◽  
Histamine Content ◽  
Plate Count ◽  
Growth Data ◽  
Tuna Fish ◽  
Effect Of Ph ◽  
Histamine Formation ◽  
Optimal Growth Rate

Abstract The present work describes the effect of pH on the growth rate and histamine formation by Klebsiella pneumoniae CK02 and Raoultella ornithinolytica TN01. Bacteria were inoculated on Tuna Fish Infusion Broth media with pH 5, 6, 7, 8 at 30°C for 6 hours. Sampling was conducted at 0, 3, and 6 hours to observe the bacteria number and calculate the histamine content formed in the medium. The number of bacteria was calculated using the Total Plate Count method, and the histamine content was analyzed using Thin Layer Chromatography with a combination of ImageJ software. Growth data and incubation time were plotted in the DMFit program to obtain growth rates. The effect of pH on growth rate and histamine formation was analyzed by ANOVA test and Duncan Multiple Range Test. The results revealed that pH affects the growth rate and histamine formation of K. pneumoniae CK02 and R. ornithinolytica TN01. The optimal growth rate of K. pneumoniae CK02 was in the range of 6-8 (0.304-0.380 log CFU/h), with the highest histamine formation ability at pH 7 (824 ppm). R. ornithinolytica TN01 had an optimal growth rate at pH 6-7 (0.480-0.508 log CFU/ml), with optimal ability to produce histamine at pH 6-8 (620-1,077.5 ppm). At pH 5, the growth rate and the ability of histamine formation by K. pneumoniae CK02 and R. ornithinolytica TN01 were inhibited.

scholarly journals Complex Regulation of Urease Formation from the Two Promoters of the ure Operon of Klebsiella pneumoniae

2007 ◽  
Vol 189 (21) ◽  
pp. 7593-7599 ◽  
Author(s):  
Qiong Liu ◽  
Robert A. Bender
Keyword(s):  
Growth Rate ◽  
Klebsiella Pneumoniae ◽  
Specific Activity ◽  
Regulatory System ◽  
Fold Increase ◽  
Sole Source ◽  
Proximal Promoter ◽  
Absolute Requirement ◽  
Activity Difference ◽  
Operon Expression

ABSTRACT Klebsiella pneumoniae can use urea as the sole source of nitrogen, thanks to a urease encoded by the ureDABCEFG operon. Expression of this operon is independent of urea and is regulated by the supply of nitrogen in the growth medium. When cells were growth rate limited for nitrogen, the specific activity of urease was about 70 times higher than that in cells grown under conditions of excess nitrogen. Much of this nitrogen regulation of urease formation depended on the nitrogen regulatory system acting through the nitrogen assimilation control protein, NAC. In a strain deleted for the nac gene, nitrogen limitation resulted in only a 7-fold increase in the specific activity of urease, in contrast to the 70-fold increase seen in that of the wild type. The ure operon was transcribed from two promoters. The proximal promoter (P1) had an absolute requirement for NAC; little or no transcription was seen in the absence of NAC. The distal promoter (P2) was independent of NAC, but its activity increased about threefold when the growth rate of the cells was limited by the nitrogen source. Transcriptional regulation of P1 and P2 accounted for most of the changes in urease activity seen under various nitrogen conditions. However, when transcription of ureDABCEFG was less than 20% of its maximum, the amount of active urease formed per transcript of ure decreased almost linearly with decreasing transcription. This may reflect a defect in the assembly of active urease and accounted for as much as a threefold activity difference under the conditions tested here. Thus, the ure operon was transcribed from a NAC-independent promoter (P2) and the most strongly NAC-dependent promoter known (P1). Most of the regulation of urease formation was transcriptional, but when ure transcription was low, assembly of active urease also was defective.

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