Time to act-assessing variations in qPCR analyses in biological nitrogen removal with examples from partial nitritation/anammox systems

Shelesh Agrawal; David G Weissbrodt; Medini Annavajhala; Marlene Mark Jensen; Jose Maria Carvajal Arroyo; George Wells; Kartik Chandran; Siegfried E Vlaeminck; Akihiko Terada; Barth F. Smets; Susanne Lackner

doi:10.1016/j.watres.2020.116604

Time to act-assessing variations in qPCR analyses in biological nitrogen removal with examples from partial nitritation/anammox systems

Shelesh Agrawal^*, David G Weissbrodt, Medini Annavajhala, Marlene Mark Jensen, Jose Maria Carvajal Arroyo, George Wells, Kartik Chandran, Siegfried E Vlaeminck, Akihiko Terada, Barth F. Smets, Susanne Lackner

^*Corresponding author for this work

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Abstract

Quantitative PCR (qPCR) is broadly used as the gold standard to quantify microbial community fractions in environmental microbiology and biotechnology. Benchmarking efforts to ensure the comparability of qPCR data for environmental bioprocesses are still scarce. Also, for partial nitritation/anammox (PN/A) systems systematic investigations are still missing, rendering meta-analysis of reported trends and generic insights potentially precarious. We report a baseline investigation of the variability of qPCR-based analyses for microbial communities applied to PN/A systems. Round-robin testing was performed for three PN/A biomass samples in six laboratories, using the respective in-house DNA extraction and qPCR protocols. The concentration of extracted DNA was significantly different between labs, ranged between 2.7 and 328 ng mg-1 wet biomass. The variability among the qPCR abundance data of different labs was very high (1-7 log fold) but differed for different target microbial guilds. DNA extraction caused maximum variation (3-7 log fold), followed by the primers (1-3 log fold). These insights will guide environmental scientists and engineers as well as treatment plant operators in the interpretation of qPCR data.

Original language	English
Article number	116604
Journal	Water Research
Volume	190
Number of pages	8
ISSN	0043-1354
DOIs	https://doi.org/10.1016/j.watres.2020.116604
Publication status	Published - 2021

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Agrawal, S., Weissbrodt, D. G., Annavajhala, M., Jensen, M. M., Arroyo, J. M. C., Wells, G., Chandran, K., Vlaeminck, S. E., Terada, A., Smets, B. F., & Lackner, S. (2021). Time to act-assessing variations in qPCR analyses in biological nitrogen removal with examples from partial nitritation/anammox systems. Water Research, 190, Article 116604. https://doi.org/10.1016/j.watres.2020.116604

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title = "Time to act-assessing variations in qPCR analyses in biological nitrogen removal with examples from partial nitritation/anammox systems",

abstract = "Quantitative PCR (qPCR) is broadly used as the gold standard to quantify microbial community fractions in environmental microbiology and biotechnology. Benchmarking efforts to ensure the comparability of qPCR data for environmental bioprocesses are still scarce. Also, for partial nitritation/anammox (PN/A) systems systematic investigations are still missing, rendering meta-analysis of reported trends and generic insights potentially precarious. We report a baseline investigation of the variability of qPCR-based analyses for microbial communities applied to PN/A systems. Round-robin testing was performed for three PN/A biomass samples in six laboratories, using the respective in-house DNA extraction and qPCR protocols. The concentration of extracted DNA was significantly different between labs, ranged between 2.7 and 328 ng mg-1 wet biomass. The variability among the qPCR abundance data of different labs was very high (1-7 log fold) but differed for different target microbial guilds. DNA extraction caused maximum variation (3-7 log fold), followed by the primers (1-3 log fold). These insights will guide environmental scientists and engineers as well as treatment plant operators in the interpretation of qPCR data.",

author = "Shelesh Agrawal and Weissbrodt, {David G} and Medini Annavajhala and Jensen, {Marlene Mark} and Arroyo, {Jose Maria Carvajal} and George Wells and Kartik Chandran and Vlaeminck, {Siegfried E} and Akihiko Terada and Smets, {Barth F.} and Susanne Lackner",

year = "2021",

doi = "10.1016/j.watres.2020.116604",

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T1 - Time to act-assessing variations in qPCR analyses in biological nitrogen removal with examples from partial nitritation/anammox systems

AU - Agrawal, Shelesh

AU - Weissbrodt, David G

AU - Annavajhala, Medini

AU - Jensen, Marlene Mark

AU - Arroyo, Jose Maria Carvajal

AU - Wells, George

AU - Chandran, Kartik

AU - Vlaeminck, Siegfried E

AU - Terada, Akihiko

AU - Smets, Barth F.

AU - Lackner, Susanne

PY - 2021

Y1 - 2021

N2 - Quantitative PCR (qPCR) is broadly used as the gold standard to quantify microbial community fractions in environmental microbiology and biotechnology. Benchmarking efforts to ensure the comparability of qPCR data for environmental bioprocesses are still scarce. Also, for partial nitritation/anammox (PN/A) systems systematic investigations are still missing, rendering meta-analysis of reported trends and generic insights potentially precarious. We report a baseline investigation of the variability of qPCR-based analyses for microbial communities applied to PN/A systems. Round-robin testing was performed for three PN/A biomass samples in six laboratories, using the respective in-house DNA extraction and qPCR protocols. The concentration of extracted DNA was significantly different between labs, ranged between 2.7 and 328 ng mg-1 wet biomass. The variability among the qPCR abundance data of different labs was very high (1-7 log fold) but differed for different target microbial guilds. DNA extraction caused maximum variation (3-7 log fold), followed by the primers (1-3 log fold). These insights will guide environmental scientists and engineers as well as treatment plant operators in the interpretation of qPCR data.

AB - Quantitative PCR (qPCR) is broadly used as the gold standard to quantify microbial community fractions in environmental microbiology and biotechnology. Benchmarking efforts to ensure the comparability of qPCR data for environmental bioprocesses are still scarce. Also, for partial nitritation/anammox (PN/A) systems systematic investigations are still missing, rendering meta-analysis of reported trends and generic insights potentially precarious. We report a baseline investigation of the variability of qPCR-based analyses for microbial communities applied to PN/A systems. Round-robin testing was performed for three PN/A biomass samples in six laboratories, using the respective in-house DNA extraction and qPCR protocols. The concentration of extracted DNA was significantly different between labs, ranged between 2.7 and 328 ng mg-1 wet biomass. The variability among the qPCR abundance data of different labs was very high (1-7 log fold) but differed for different target microbial guilds. DNA extraction caused maximum variation (3-7 log fold), followed by the primers (1-3 log fold). These insights will guide environmental scientists and engineers as well as treatment plant operators in the interpretation of qPCR data.

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DO - 10.1016/j.watres.2020.116604

M3 - Journal article

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SN - 0043-1354

VL - 190

JO - Water Research

JF - Water Research

M1 - 116604

ER -

Time to act-assessing variations in qPCR analyses in biological nitrogen removal with examples from partial nitritation/anammox systems

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