Determination of<i>Pseudomonas Putida</i>Live Cells with Classic Cultivation and Staining with “Live/Dead Baclight Bacterial Viability Kit”

Ivanova, Iliana; Kambarev, Stanimir; Popova, R. A.; Naumovska, E.G.; Markoska, K.; Dushkin, Ceco D.

doi:10.1080/13102818.2010.10817898

Cited by 12 publications

(5 citation statements)

References 8 publications

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“…After the EPS extraction, the remaining algae were collected and regarded as the algae without EPS. The TEM characterization, Live/Dead test 62 , and 96 h growth assay were performed to assess the potential effect of the EPS extraction on the algae. As one of the major components of EPS, the secretion kinetics of extracellular protein from the untreated and EPS-extracted algal cells in the absence and presence of the AgNPs and Ag + (from AgNO 3 ) were also determined.…”

Section: Methodsmentioning

confidence: 99%

The role of exopolymeric substances in the bioaccumulation and toxicity of Ag nanoparticles to algae

Zhou

Zhang

et al. 2016

Sci Rep

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Exopolymeric substances (EPS) have an important role in bioaccumulation and toxicity of nanoparticles (NPs) to algae, which warrants specific studies. The interaction of EPS with citrate and polyvinyl pyrrolidone (PVP) coated AgNPs (C-AgNPs and P-AgNPs, respectively) and its roles in bioaccumulation and toxicity of the AgNPs to Chlorella pyrenoidosa were investigated. The amino and aromatic carboxylic groups in the EPS were involved in the EPS-AgNP interactions. Compared with Ag+, C-AgNPs had comparable total bioaccumulation but greater absorption by intact algae with EPS; P-AgNPs had the smallest total bioaccumulation and were mainly adsorbed on algal surfaces. With EPS removed, the total bioaccumulations and surface adsorptions for the three Ag species decreased but the cell internalizations increased; the 96 h half growth inhibition concentrations decreased, indicating EPS alleviated the algal toxicity of Ag. The cell-internalized but not the adsorbed AgNPs could contribute to the nanotoxicity. The EPS could bind both AgNPs and Ag+, and thus inhibited the cell internalization and the nanotoxicity. However, the EPS-bound Ag on the cell surfaces would migrate along with the algae and be biologically amplified in the aquatic food chains, presenting ecological risks. These results are helpful for understanding the fate and ecological effects of NPs.

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Section: Methodsmentioning

confidence: 99%

The role of exopolymeric substances in the bioaccumulation and toxicity of Ag nanoparticles to algae

Zhou

Zhang

et al. 2016

Sci Rep

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“…After cultivation on solid rich medium, these cells give the slow growing colonies, which are visible after 48 h. These facts suggest that some of the red cells, which have been marked as dead, are viable and could restore their normal functions, giving colonies that emerge later. We proved the possibility of some sub lethal membrane damages which could bring to red staining and confusion in the results interpretation [31].…”

Section: The Behavior Of Pseudomonas Putida Cells On Thin Zno Film Is Verymentioning

confidence: 81%

Effect of ZnO Thin Films on Survival of Pseudomonas Cells

Ia¹,

Tasheva‐Terzieva²,

Angelov³

et al. 2012

J Nanomedic Nanotechnol

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Thin films of ZnO nanoparticles obtained by magnetron sputtering deposition and wet chemical methods are investigated for their antibacterial effect on Gracilicutes-bacteria Pseudomonas putida. This bacterium is used as a sensitive standard in an international water quality test, ISO 10712:1995. It was chosen for toxic assessment of thin ZnO films, prepared by magnetron sputtering, sol-gel dip coating and chemical bath deposition methods. Washed and stained with Dead/Live bacterial kit suspension is evenly distributed on thin ZnO films and the survived cells are counted every 2 hours. Two microbiological methods are used: direct counting with epifluorescent microscope Leika DM 5500B and cultivation in nutrient medium. The results are processed with different statistical methods and show bactericidal effect of thin ZnO films. The surface relief of thin nanostructured film is crucial for the antibacterial effect and the fastest bactericidal effect demonstrates the ZnO film obtained by sol-gel dip coating. Journal of Nanomedicine & NanotechnologyJ o u rna l of N a n o m ed icine & N a n o te chnolo g y

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“…It was reported that viable P . putida (ATCC 12633) cells were stained by propidium iodide suggesting that the use of LIVE/DEAD BacLight Bacterial Viability kit may give a confusing result in determining live cells [50]. We propose that additional studies like TEM, changes in membrane fluidity, changes in membrane potential and changes in membrane phospholipid composition will be required to clarify the level of membrane damage in our viable but non-culturable bacterial cells subjected to desiccation-rehydration process.…”

Section: Discussionmentioning

confidence: 99%

Desiccation-induced viable but nonculturable state in Pseudomonas putida KT2440, a survival strategy

et al. 2019

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The potential of Pseudomonas putida KT2440 to act as a plant-growth promoter or as a bioremediator of toxic compounds can be affected by desiccation. In the present work, the bacterial survival ratio (BSR) in response to air desiccation was evaluated for P . putida KT2440 in the presence of different protectors. The BSR in the presence of nonreducing disaccharides, such as trehalose, was high after 15 days of desiccation stress (occurring at 30°C and 50% relative humidity), whereas in the absence of a protector the bacterial counts diminished to nondetectable numbers (ca 2.8 log CFU/mL). The LIVE/DEAD staining method showed that bacteria protected with trehalose maintained increased numbers of green cells after desiccation while cells without protection were all observed to be red. This indicated that nonprotected bacteria had compromised membrane integrity. However, when nonprotected bacteria subjected to 18 days of desiccation stress were rehydrated for a short time with maize root exudates or for 48 h with water (prolonged rehydration), the bacterial counts were as high as that observed for those not subjected to desiccation stress, suggesting that the cells entered the viable but nonculturable (VBNC) state under desiccation and that they returned to a culturable state after those means of rehydration. Interestingly an increase in the green color intensity of cells that returned to a culturable state was observed using LIVE/DEAD staining method, indicating an improvement in their membrane integrity. Cellular activity in the VBNC state was determined. A GFP-tagged P . putida strain expressing GFP constitutively was subjected to desiccation. After 12 days of desiccation, the GFP-tagged strain lost culturability, but it exhibited active GFP expression, which in turn made the cells green. Furthermore, the expression of 16S rRNA, rpoN (housekeeping), mutL , mutS (encoding proteins from the mismatch repair complex), and oprH (encoding an outer membrane protein) were examined by RT-PCR. All evaluated genes were expressed by both types of cells, culturable and nonculturable, indicating active molecular processes during the VBNC state.

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Determination ofPseudomonas PutidaLive Cells with Classic Cultivation and Staining with “Live/Dead Baclight Bacterial Viability Kit”

Abstract: ABSTRACT

Cited by 12 publications

References 8 publications

The role of exopolymeric substances in the bioaccumulation and toxicity of Ag nanoparticles to algae

The role of exopolymeric substances in the bioaccumulation and toxicity of Ag nanoparticles to algae

Effect of ZnO Thin Films on Survival of Pseudomonas Cells

Desiccation-induced viable but nonculturable state in Pseudomonas putida KT2440, a survival strategy

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