scholarly journals Probing the viability of palladium-challenged bacterial cells using flow cytometry

2018 ◽  
Vol 94 (1) ◽  
pp. 295-301 ◽  
Author(s):  
Jacob B Omajali ◽  
Iryna P Mikheenko ◽  
Tim W Overton ◽  
Mohamed L Merroun ◽  
Lynne E Macaskie
Keyword(s):  
Flow Cytometry ◽  
Bacterial Cells

scholarly journals Using flow cytometry and light-induced fluorescence technique to characterizethe variability and characteristics of bioaerosols in springtime at Metro Atlanta, Georgia

2018 ◽  
Author(s):  
Arnaldo Negron ◽  
Natasha DeLeon-Rodriguez ◽  
Samantha M. Waters ◽  
Luke D. Ziemba ◽  
Bruce Anderson ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Nucleic Acid ◽  
Acid Content ◽  
Fungal Spores ◽  
Epifluorescence Microscopy ◽  
Nucleic Acid Content ◽  
Rain Event ◽  
Bacterial Cells ◽  
Sampling System ◽  
Spores And Pollen

Abstract. The abundance and speciation of primary biological aerosol particles (PBAP) is important for understanding their impacts on human health, cloud formation and ecosystems. Towards this, we have developed a protocol for quantifying PBAP collected from large volumes of air with a portable wet-walled cyclone bioaerosol sampler. A flow cytometry (FCM) protocol was then developed to quantify and characterize the PBAP populations from the sampler, which were confirmed against epifluorescence microscopy. The sampling system and FCM analysis were used to study PBAP in Atlanta, GA over a two-month period and showed clearly defined populations of DNA-containing particles: Low Nucleic Acid-content particles (bioLNA), High Nucleic Acid-content particles (HNA) being fungal spores and pollen. We find that daily-average springtime PBAP concentration (1 to 5 μm diameter) ranged between 1.4 × 104 and 1.1 × 105 m−3. The BioLNA population dominated PBAP during dry days (72 ± 18 %); HNA dominated the PBAP during humid days and following rain events, where HNA (e.g., wet-ejected fungal spores) comprised up to 92 % of the PBAP number. Concurrent measurements with a Wideband Integrated Bioaerosol Sensor (WIBS-4A) showed that FBAP and total FCM counts are similar; HNA (from FCM) significantly correlated with ABC type FBAP concentrations throughout the sampling period (and for the same particle size range, 1–5 μm diameter). However, the FCM bioLNA population, possibly containing bacterial cells, did not correlate to any FBAP type. The lack of correlation of any WIBS FBAP type with the bioLNA suggest bacterial cells may be more difficult to detect with autofluorescence than previously thought. Ιdentification of bacterial cells even in the FCM (bioLNA population) is challenging, given that the fluorescence level of stained cells at times may be comparable to that seen from abiotic particles. HNA and ABC displayed highest concentration on a humid and warm day after a rain event (4/14), suggesting that both populations correspond to wet-ejected fungal spores. Overall, information from both instruments combined reveals a highly dynamic airborne bioaerosol community over Atlanta, with a considerable presence of fungal spores during humid days, and a bioLNA population dominating bioaerosol community during dry days.

Preliminary Study on In Situ Realtime Quantitation of Target Bacteria on the Principle of Flow Cytometry

2017 ◽  
Vol 262 ◽  
pp. 224-227
Author(s):  
Gen Murakami ◽  
Yuichi Sugai ◽  
Kyuro Sasaki
Keyword(s):  
Flow Cytometry ◽  
Bacterial Culture ◽  
Scattered Light ◽  
Bacterial Suspension ◽  
Culture Solution ◽  
Bacterial Cells ◽  
Monitoring Wells ◽  
Measurement Principle ◽  
Preliminary Study

In-situ realtime method that can monitor the target bacteria should be used to determine the real situation of the bacteria in deep parts of heaps in heap bioleaching plants. This study suggest to apply flow cytometry technology to in-situ realtime monitoring of target bacteria. Flow cytometry is a method that can rapidly quantify the bacterial cells in bacterial suspension based on the detection of lights that are emitted from bacterial cells. In this study, we estimated the possibility of the application of flow cytometry to the selective detection of target bacteria. The bacterial culture solution that had been diluted by water including other bacteria was provided for fluorescence spectral analysis and scattered light analysis that were functions of flow cytometry. Our target bacteria could be selectively detected by those analyses in this study, therefore, it was shown that the flow cytometry could be useful for detecting target bacteria selectively. Because the measurement principle of flow cytometry is quite simple, it can be expected to be installed into deep heaps through the monitoring wells and determine the dominance of target bacteria in-situ and realtime in the future.

scholarly journals THE INFLUENCE OF ELECTROMAGNETIC FIELD ON MORPHOPHYSIOLOGICAL PARAMETERS OF BACTERIAL CELLS EVALUATED THROUGH FLOW CYTOMETRY

2016 ◽  
Author(s):  
Elena Radu ◽  
◽  
F. Marinescu ◽  
I. Savin ◽  
C. M. Kamerzan (Saviuc) ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Electromagnetic Field ◽  
Bacterial Cells ◽  
Through Flow

scholarly journals Population Structure and Morphotype Analysis of “CandidatusAccumulibacter” Using FluorescenceIn SituHybridization-Staining-Flow Cytometry

2019 ◽  
Vol 85 (9) ◽  
Author(s):  
Chao Li ◽  
Wei Zeng ◽  
Ning Li ◽  
Yu Guo ◽  
Yongzhen Peng
Keyword(s):  
Flow Cytometry ◽  
Population Structure ◽  
Electron Acceptor ◽  
Phosphorus Removal ◽  
Fish Analysis ◽  
Bacterial Cells ◽  
Denitrifying Phosphorus Removal ◽  
Optimal Method ◽  
Total Bacteria

ABSTRACT“CandidatusAccumulibacter” is the dominant polyphosphate-accumulating organism (PAO) in denitrifying phosphorus removal (DPR) systems. In order to investigate the community structure and clade morphotypes of “CandidatusAccumulibacter” in DPR systems through flow cytometry (FCM), denitrifying phosphorus removal of almost 100% using nitrite and nitrate as the electron acceptor was achieved in sequencing batch reactors (SBRs). An optimal method of flow cytometry combined with fluorescencein situhybridization and SYBR green I staining (FISH-staining-flow cytometry) was developed to quantify PAOs in DPR systems. By setting the width value of FCM, bacterial cells in a sludge sample were divided into three groups in different morphotypes, namely, coccus, coccobacillus, and bacillus. Average percentages that the three different PAO populations accounted for among total bacteria from SBR1 (SBR2) were 42% (45%), 14% (13%), and 4% (2%). FCM showed that the ratios of PAOs to total bacteria in the two reactors were 61% and 59%, and the quantitative PCR (qPCR) results indicated that IIC was the dominant “CandidatusAccumulibacter” clade in both denitrifying phosphorus removal systems, reaching 50% of the total “CandidatusAccumulibacter” bacteria. The subdominant clade in the reactor with nitrite as the electron acceptor was IID, accounting for 31% of the total “CandidatusAccumulibacter” bacteria. The FCM and qPCR results suggested that clades IIC and IID were both coccus, clade IIF was coccobacillus, and clade IA was bacillus. FISH analysis also indicated that PAOs were major cocci in the systems. An equivalence test of FCM-based quantification confirmed the accuracy of FISH-staining-flow cytometry, which can meet the quantitative requirements for PAOs in complex activated sludge samples.IMPORTANCEAs one group of the most important functional phosphorus removal organisms, “CandidatusAccumulibacter,” affiliated with theRhodocyclusgroup of theBetaproteobacteria, is a widely recognized and studied PAO in the field of biological wastewater treatment. The morphotypes and population structure of clade-level “CandidatusAccumulibacter” were studied through novel FISH-staining-flow cytometry, which involved denitrifying phosphorus removal (DPR) achieving carbon and energy savings and simultaneous removal of N and P, thus inferring the different denitrifying phosphorus removal abilities of these clades. Additionally, based on this method,in situquantification for specific polyphosphate-accumulating organisms (PAOs) enables a more efficient process and more accurate result. The establishment of FISH-staining-flow cytometry makes cell sorting of clade-level noncultivated organisms available.

scholarly journals Determination of Total Protein Content of Bacterial Cells by SYPRO Staining and Flow Cytometry

1999 ◽  
Vol 65 (7) ◽  
pp. 3251-3257 ◽  
Author(s):  
Mikhail V. Zubkov ◽  
Bernhard M. Fuchs ◽  
Heike Eilers ◽  
Peter H. Burkill ◽  
Rudolf Amann
Keyword(s):  
Flow Cytometry ◽  
Protein Content ◽  
Total Protein ◽  
Total Protein Content ◽  
Bacterial Cells ◽  
Protein Fluorescence ◽  
The North ◽  
Planktonic Bacteria ◽  
Different Temperatures ◽  
Protein Cell

ABSTRACT An assay has been developed for measuring protein biomass of marine planktonic bacteria by flow cytometry. The method was calibrated by using five species of Bacteria (an Arcobactersp., a Cytophaga sp., an Oceanospirillum sp., aPseudoalteromonas sp., and a Vibrio sp.) recently isolated from seawater samples and grown in culture at different temperatures. The intensity of SYPRO-protein fluorescence of these bacteria strongly correlated with their total protein content, measured by the bicinchoninic acid method to be in the range of 60 to 330 fg of protein cell−1(r 2 = 0.93, n = 34). According to the calibration, the mean biomass of planktonic bacteria from the North Sea in August 1998 was 24 fg of protein cell−1.

scholarly journals Specific and Rapid Enumeration of Viable but Nonculturable and Viable-Culturable Gram-Negative Bacteria by Using Flow Cytometry

2010 ◽  
Vol 76 (15) ◽  
pp. 5088-5096 ◽  
Author(s):  
Mohiuddin M. Taimur Khan ◽  
Barry H. Pyle ◽  
Anne K. Camper
Keyword(s):  
Flow Cytometry ◽  
Pseudomonas Syringae ◽  
Phase 1 ◽  
Epifluorescence Microscopy ◽  
Gram Negative Bacteria ◽  
Bacterial Cells ◽  
Culture Methods ◽  
Viable But Nonculturable ◽  
Gram Negative ◽  
Analytical Technique

ABSTRACT An issue of critical concern in microbiology is the ability to detect viable but nonculturable (VBNC) and viable-culturable (VC) cells by methods other than existing approaches. Culture methods are selective and underestimate the real population, and other options (direct viable count and the double-staining method using epifluorescence microscopy and inhibitory substance-influenced molecular methods) are also biased and time-consuming. A rapid approach that reduces selectivity, decreases bias from sample storage and incubation, and reduces assay time is needed. Flow cytometry is a sensitive analytical technique that can rapidly monitor physiological states of bacteria. This report outlines a method to optimize staining protocols and the flow cytometer (FCM) instrument settings for the enumeration of VBNC and VC bacterial cells within 70 min. Experiments were performed using the FCM to quantify VBNC and VC Escherichia coli O157:H7, Pseudomonas aeruginosa, Pseudomonas syringae, and Salmonella enterica serovar Typhimurium cells after staining with different fluorescent probes: SYTO 9, SYTO 13, SYTO 17, SYTO 40, and propidium iodide (PI). The FCM data were compared with those for specific standard nutrient agar to enumerate the number of cells in different states. By comparing results from cultures at late log phase, 1 to 64% of cells were nonculturable, 40 to 98% were culturable, and 0.7 to 4.5% had damaged cell membranes and were therefore theoretically dead. Data obtained using four different Gram-negative bacteria exposed to heat and stained with PI also illustrate the usefulness of the approach for the rapid and unbiased detection of dead versus live organisms.

scholarly journals Application of gel microdroplet and flow cytometry techniques to selective enrichment of non-growing bacterial cells

2001 ◽  
Vol 197 (1) ◽  
pp. 29-33 ◽  
Author(s):  
Akira Manome ◽  
Hui Zhang ◽  
Yoshiki Tani ◽  
Tohoru Katsuragi ◽  
Ryuichiro Kurane ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Bacterial Cells ◽  
Selective Enrichment

scholarly journals Determining the Development of Persisters in Extensively Drug-Resistant Acinetobacter baumannii upon Exposure to Polymyxin B-Based Antibiotic Combinations Using Flow Cytometry

2019 ◽  
Vol 64 (3) ◽  
Author(s):  
Fiona Hui-Sian Wong ◽  
Yiying Cai ◽  
Hui Leck ◽  
Tze-Peng Lim ◽  
Jocelyn Qi-Min Teo ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Acinetobacter Baumannii ◽  
Polymyxin B ◽  
Live Cells ◽  
Drug Resistant ◽  
Bacterial Cells ◽  
Persister Cells ◽  
Content Type ◽  
Initial Inoculum ◽  
Extensively Drug Resistant

ABSTRACT Polymyxin B-based combinations are increasingly prescribed as a last-line option against extensively drug-resistant (XDR) Acinetobacter baumannii. It is unknown if such combinations can result in the development of nondividing persister cells in XDR A. baumannii. We investigated persister development upon exposure of XDR A. baumannii to polymyxin B-based antibiotic combinations using flow cytometry. Time-kill studies (TKSs) were conducted in three nonclonal XDR A. baumannii strains with 5 log10 CFU/ml bacteria against polymyxin B alone and polymyxin B-based two-drug combinations over 24 h. At different time points, samples were obtained and enumerated by viable plating and flow cytometry. Propidium iodide and carboxyfluorescein succinimidyl ester dyes were used to differentiate between live and dead cells and between dividing and nondividing cells, respectively, at the single-cell level, and nondividing live cells were resuscitated and characterized phenotypically. Our results from viable plating showed that polymyxin B plus meropenem and polymyxin B plus rifampin were each bactericidal (>99.9% kill compared to the initial inoculum) against 2/3 XDR A. baumannii strains at 24 h. By flow cytometry, however, none of the combinations were bactericidal against XDR A. baumannii at 24 h. Further analysis using cellular dyes in flow cytometry revealed that upon exposure to polymyxin B-based combinations, XDR A. baumannii entered a viable but nondividing persister state. These bacterial cells reinitiated division upon the removal of antibiotic pressure and did not have a growth deficit compared to the parent strain. We conclude that persister cells develop in XDR A. baumannii upon exposure to polymyxin B-based combinations and that nonplating methods appear to complement viable-plating methods in describing the killing activity of polymyxin B-based combinations against XDR A. baumannii.

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