Quantifying synthetic bacterial community composition with flow cytometry: efficacy in mock communities and challenges in co-cultures DOI Creative Commons
Fabian Mermans, Ioanna Chatzigiannidou, Wim Teughels

et al.

mSystems, Journal Year: 2024, Volume and Issue: unknown

Published: Nov. 29, 2024

ABSTRACT Determination of bacterial community composition in synthetic communities is critical for understanding microbial systems. The typically determined through plating or PCR-based methods, which can be labor-intensive, expensive, prone to bias. Simultaneously, flow cytometry has been suggested as a cheap and fast alternative. However, since the technique captures phenotypic state cells, accurate determination could affected when bacteria are co-cultured. We investigated performance quantifying oral compared it strain specific qPCR 16S rRNA gene amplicon sequencing. Therefore, axenic cultures, mock co-cultures were prepared. Random forest classifiers trained on data cultures used determine communities, well Flow was shown have lower average root mean squared error outperformed methods even (flow cytometry: 0.11 ± 0.04; qPCR: 0.26 0.09; sequencing: 0.15 0.01). When co-cultured, neither cytometry, strain-specific qPCR, nor sequencing resulted similar composition. Performance decreased with due changing phenotypes. Finally, discrepancies between found. These findings highlight challenges ahead by cytometry. IMPORTANCE Quantification crucial steering interactions. Traditional approaches like plating, often labor-intensive expensive limit high-throughput experiments. Recently, swift alternative successfully demonstrated simple communities. measures measurements differing Especially, phenotypes resulting from co-culturing profound effect applicability this context. This research illustrates feasibility structure co-cultures.

Language: Английский

Full-electric microfluidic platform to capture, analyze and selectively release single cells DOI Creative Commons
Ruben Van den Eeckhoudt, An‐Sofie Christiaens, Frederik Ceyssens

et al.

Lab on a Chip, Journal Year: 2023, Volume and Issue: 23(19), P. 4276 - 4286

Published: Jan. 1, 2023

Current single-cell technologies require large and expensive equipment, limiting their use to specialized labs. In this paper, we present for the first time a microfluidic device which demonstrates combined method full-electric cell capturing, analyzing, selectively releasing with resolution. All functionalities are experimentally demonstrated on Saccharomyces cerevisiae. Our platform consists of traps centered around pair individually accessible coplanar electrodes, positioned under channel. Using device, validate our novel Two-Voltage trapping single cells by positive dielectrophoresis (pDEP). Cells attracted trap when high voltage (VH) is applied. A low (VL) holds already trapped in place without attracting additional cells, allowing full control over number cells. After trapping, analyzed broadband electrochemical impedance spectroscopy. These measurements allow detection extraction parameters. Additionally, these show strong correlation between average phase change size, enabling system size biological applications. Finally, allows turning off pDEP signal trap. The experimental results techniques potential as analysis tool miniaturization automation opens new avenues towards small-scale, throughput sorting lab-on-CMOS devices.

Language: Английский

Citations

5
Raman cell sorting for single-cell research DOI Creative Commons
Xusheng Tang, Qingyi Wu, Lindong Shang

et al.

Frontiers in Bioengineering and Biotechnology, Journal Year: 2024, Volume and Issue: 12

Published: May 20, 2024

Cells constitute the fundamental units of living organisms. Investigating individual differences at single-cell level facilitates an understanding cell differentiation, development, gene expression, and cellular characteristics, unveiling underlying laws governing life activities in depth. In recent years, integration manipulation recognition technologies into detection sorting systems has emerged as a powerful tool for advancing research. Raman technology garnered attention owing to its non-labeling, non-destructive features capability analyze samples containing water. addition, this can provide live cells subsequent genomics analysis sequencing. This paper emphasizes importance research, describes research methods that currently exist, including identification techniques, highlights advantages spectroscopy field by comparing it with fluorescence-activated (FACS) technique. It various existing techniques introduces their respective disadvantages. The above were compared analyzed, considering variety factors. current bottlenecks include weak spontaneous signals requirement prolonged total exposure time, significantly constraining technology's speed, efficiency, throughput. provides overview enhancing associated Finally, outlines detailed information related mentioned discusses development trends direction sorting.

Language: Английский

Citations

1
Quantifying synthetic bacterial community composition with flow cytometry: efficacy in mock communities and challenges in co-cultures DOI Open Access
Fabian Mermans, Ioanna Chatzigiannidou, Wim Teughels

et al.

bioRxiv (Cold Spring Harbor Laboratory), Journal Year: 2024, Volume and Issue: unknown

Published: July 26, 2024

Abstract Determination of bacterial community composition in synthetic communities is critical for understanding microbial systems. The typically determined through plating or PCR-based methods which can be labor-intensive, expensive prone to bias. Simultaneously, flow cytometry has been suggested as a cheap and fast alternative. However, since the technique captures phenotypic state cells, accurate determination could affected when bacteria are co-cultured. We investigated performance quantifying oral compared it strain specific qPCR 16S rRNA gene amplicon sequencing. Therefore, axenic cultures, mock co-cultures were prepared. Random forest classifiers trained on data cultures used determine communities, well Flow was shown have lower average root mean squared error outperformed even (flow cytometry: 0.11 ± 0.04; qPCR: 0.26 0.09; sequencing: 0.15 0.01). When co-cultured, neither cytometry, sequencing resulted similar composition. Performance decreased due changing phenotypes. Finally, discrepancies between found. These findings highlight challenges ahead by cytometry. Importance Quantification crucial steering interactions. Traditional approaches like plating, often labor-intensive limit high-throughput experiments. Recently, swift alternative successfully demonstrated simple communities. measures measurements differing Especially phenotypes resulting from co-culturing profound effect applicability this context. This research illustrates feasibility structure co-cultures.

Language: Английский

Citations

1
Microbial Biotechnology: Pioneering Ecological Innovations for a Sustainable Future DOI Open Access
Murali Krishna Pasupuleti

Published: Aug. 11, 2024

Abstract: This chapter explores the transformative potential of microbial biotechnology in addressing some most pressing ecological challenges our time. It delves into diverse applications microbes environmental sustainability, agriculture, and industry, highlighting their role processes such as bioremediation, biofuel production, sustainable agriculture. The also examines latest advances genomics synthetic biology, showcasing how these technologies are driving innovation enabling development novel solutions for climate change mitigation, resource recovery, pollution control. Ethical considerations associated with use discussed, alongside a forward-looking vision innovations can contribute to more resilient future. emphasizes importance interdisciplinary collaboration global commitment harnessing power achieve sustainability address challenges. Keywords: Microbial Biotechnology, Environmental Sustainability, Bioremediation, Biofuels, Sustainable Agriculture, Genomics, Synthetic Biology, Carbon Sequestration, Circular Economy, Ecological Innovation.

Language: Английский

Citations

0
Quantifying synthetic bacterial community composition with flow cytometry: efficacy in mock communities and challenges in co-cultures DOI Creative Commons
Fabian Mermans, Ioanna Chatzigiannidou, Wim Teughels

et al.

mSystems, Journal Year: 2024, Volume and Issue: unknown

Published: Nov. 29, 2024

ABSTRACT Determination of bacterial community composition in synthetic communities is critical for understanding microbial systems. The typically determined through plating or PCR-based methods, which can be labor-intensive, expensive, prone to bias. Simultaneously, flow cytometry has been suggested as a cheap and fast alternative. However, since the technique captures phenotypic state cells, accurate determination could affected when bacteria are co-cultured. We investigated performance quantifying oral compared it strain specific qPCR 16S rRNA gene amplicon sequencing. Therefore, axenic cultures, mock co-cultures were prepared. Random forest classifiers trained on data cultures used determine communities, well Flow was shown have lower average root mean squared error outperformed methods even (flow cytometry: 0.11 ± 0.04; qPCR: 0.26 0.09; sequencing: 0.15 0.01). When co-cultured, neither cytometry, strain-specific qPCR, nor sequencing resulted similar composition. Performance decreased with due changing phenotypes. Finally, discrepancies between found. These findings highlight challenges ahead by cytometry. IMPORTANCE Quantification crucial steering interactions. Traditional approaches like plating, often labor-intensive expensive limit high-throughput experiments. Recently, swift alternative successfully demonstrated simple communities. measures measurements differing Especially, phenotypes resulting from co-culturing profound effect applicability this context. This research illustrates feasibility structure co-cultures.

Language: Английский

Citations

0