Determination of antibiotic susceptibility of bacteria by flow cytometric method

Hatipoglu, Huseyin; Erman, Gülay; Toptan, Hande; Köroğlu, Mehmet; Altındiş, Mustafa

doi:10.1007/s11274-022-03332-2

Cited by 5 publications

(6 citation statements)

References 23 publications

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“…For the routine use of FCM in the detection of bacterial contamination in PS, it would be appropriate to take a sample with a satellite bag after the PS is taken and leave it to incubate at 37 °C for 5 days like a blood culture bottle. In line with the data we obtained in our other study (Hatipoglu et al 2022 ) and as suggested by some researchers, in cases where there is no satellite bag, we think that pre-incubation of the sample taken from the PS for 1–2 h in an incubator at 37 °C would be useful in detecting the presence of bacteria with FCM. We recommend that researchers consider this process in their study design.…”

Section: Discussionsupporting

confidence: 92%

“…For bacteriological analyzes, we used the FCM analysis protocol that we used in our previous study (Hatipoglu et al 2022 ). As described in detail above, 1 ml was taken from all samples (spiked and non-spiked APSs), platelets were lysed, centrifuged, pellet suspension was prepared, fluorescent staining was performed and analyzed in a flow cytometry device (Accuri C6, Becton Dickinson, USA).…”

Section: Methodsmentioning

confidence: 99%

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Use of flow cytometry method to detect contaminations of platelet suspensions

Bolat,

Hatipoğlu,

Köroğlu

et al. 2024

World J Microbiol Biotechnol

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In this study, it was aimed to investigate bacterial contamination in apheresis platelet suspensions (APS) by automated blood culture system and flow cytometry method (FCM).33 spiked APS each using 11 bacterial strains (5 standard strains, 6 clinical isolates), were prepared in three different dilutions (1–10, 10–50, 50-100 cfu/mL), incubated in two different temperatures (35–37 °C and 22–24 °C) and different incubation times (18–96 h) evaluated by FCM. This three different dilutions were also inoculated into special platelet culture bottles (BacT/ALERT® BPA) and loaded into the blood culture system. Additionally 80 APSs routinely prepared in the Transfusion Center were evaluated by both FCM and the blood culture system. Platelets were lysed by freeze-thaw method.All spiked samples were positive with BacT/ALERT® BPA in 12–18 h. In 96 h incubation at 22–24 °C, the presence of bacteria was detected by FCM in all other samples (31/33) except low dilutions (1–10 and 10–100 CFU/ml) of K.pneumoniae standard strain. In the 35–37 °C, the presence of bacteria was detected by FCM in all samples (33/33) after 48 h of incubation. In routine APS one sample detected as positive (Bacillus simplex) with BacT/ALERT® BPA and no positivity was detected by FCM.The freeze-thaw method, which we have optimized for the lysis of platelets, is very practical and can be easily applied. The BacT/ALERT® system has been found to be very sensitive in detecting bacterial contamination in PSs. Flow cytometry method has been found to be successful, fast, easy to use and low cost in detecting bacterial contamination in PSs.

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Section: Discussionsupporting

confidence: 92%

Section: Methodsmentioning

confidence: 99%

Use of flow cytometry method to detect contaminations of platelet suspensions

Bolat,

Hatipoğlu,

Köroğlu

et al. 2024

World J Microbiol Biotechnol

Self Cite

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“…FCM assay also offers an advantage in higher flexibility compared to current automated AST tools, such as Vitek-2 [22,36]. High correlations (>90%) of flow cytometric determination of AST against Vitek-2 was documented across multiple species [22]. These flexibilities include testing any bacteria species against any antibiotics, including antibiotics combinations [37][38][39].…”

Section: Discussionmentioning

confidence: 99%

“…One major advantage of FCM is the rapid assessments of cells at the single cell resolution. FCM assay also offers an advantage in higher flexibility compared to current automated AST tools, such as Vitek-2 [ 22 , 36 ]. High correlations (>90%) of flow cytometric determination of AST against Vitek-2 was documented across multiple species [ 22 ].…”

Section: Discussionmentioning

confidence: 99%

“…Currently, there is limited literature establishing a FCM assay that can be adopted for rapid AST covering a wide range of antibiotics [19][20][21]. Flow cytometry has been demonstrated to have good correlation to current automated diagnostic methods [22]. To the best of our knowledge, flow cytometric AST has yet to be established for routine clinical use globally.…”

Section: Introductionmentioning

confidence: 99%

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Can flow cytometric measurements of reactive oxygen species levels determine minimal inhibitory concentrations and antibiotic susceptibility testing for Acinetobacter baumannii?

Yeo,

Low,

Begam

et al. 2024

PLoS ONE

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Current antimicrobial susceptibility testing (AST) requires 16–24 hours, delaying initiation of appropriate antibiotics. Hence, there is a need for rapid AST. This study aims to develop and evaluate the feasibility of a rapid flow cytometric AST assay to determine minimum inhibitory concentration (MIC) for carbapenem-resistant Acinetobacter baumannii (CRAB). Antibiotic exposure causes increased intracellular reactive oxygen species (ROS) in bacteria. We hypothesized that ROS can be used as a marker to determine MIC. We assessed three CRAB clinical isolates across fifteen antibiotics at various concentrations in a customized 96-well microtiter plate. The antibiotics assessed include amikacin, beta-lactams (ampicillin/sulbactam, aztreonam, cefepime, ceftolozane/tazobactam, doripenem, imipenem, meropenem, and piperacillin/tazobactam), levofloxacin, polymyxin B, rifampicin, trimethoprim/sulfamethoxazole, and tetracyclines (tigecycline and minocycline). These clinical CRAB isolates were assessed for ROS after antibiotic treatment. Increased ROS levels indicated by increased RedoxSensorTM Green (RSG) fluorescence intensity was assessed using flow cytometry (FCM). MIC was set as the lowest antibiotic concentration that gives a ≥1.5-fold increase in mode RSG fluorescence intensity (MICRSG). Accuracy of MICRSG was determined by comparing against microtiter broth dilution method performed under CLSI guidelines. ROS was deemed accurate in determining the MICs for β-lactams (83.3% accuracy) and trimethoprim/sulfamethoxazole (100% accuracy). In contrast, ROS is less accurate in determining MICs for levofloxacin (33.3% accuracy), rifampicin (0% accuracy), amikacin (33.3% accuracy), and tetracyclines (33.3% accuracy). Collectively, this study described an FCM-AST assay to determine antibiotic susceptibility of CRAB isolates within 5 hours, reducing turnaround time up to 19 hours.

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Antibiotic Resistance Detection in Pseudomonas aeruginosa: Recent Strategies, Advances, and Challenges

Clarindo Lopes,

Koushanpour,

Wiebe

et al. 2024

Analysis & Sensing

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Antibiotic resistance presents a worldwide public health emergency, impeding the effective management of infectious diseases. Pseudomonas aeruginosa plays a substantial role as a bacterial pathogen, particularly in infections among hospitalized individuals, and those with weakened immune systems. Timely and accurate detection of antimicrobial resistance in P. aeruginosa is crucial for initiating tailored antibiotic therapy promptly, thus improving patient outcomes. Nevertheless, this endeavor encounters challenges due to the intricate and varied nature of antibiotic‐resistant strains. Extensive efforts have been invested in developing sensors and instrumentations for assessing antibiotic resistance or susceptibility (AST), aiming to enable personalized patient treatment with appropriate medications. This paper focuses on recent advancements in these methodologies, utilized for evaluating the degree of antibiotic resistance or susceptibility inpathogenic bacteria. Flow cytometry, dual molecular recognition and mass spectrometry are presented as newer phenotypic AST techniques. Genomic methods and the pyocyanin detection are listed as methodologies for the detection of resistance indicators.

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Determination of antibiotic susceptibility of bacteria by flow cytometric method

Cited by 5 publications

References 23 publications

Use of flow cytometry method to detect contaminations of platelet suspensions

Use of flow cytometry method to detect contaminations of platelet suspensions

Can flow cytometric measurements of reactive oxygen species levels determine minimal inhibitory concentrations and antibiotic susceptibility testing for Acinetobacter baumannii?

Antibiotic Resistance Detection in Pseudomonas aeruginosa: Recent Strategies, Advances, and Challenges

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