A rapid flow cytometry method to assess bacterial abundance in agricultural soil

Bressan, Mélanie; Gattin, Isabelle; Desaire, Sylvie; Castel, Lisa; Gangneux, Christophe; Laval, Karine

doi:10.1016/j.apsoil.2014.12.007

Cited by 41 publications

(35 citation statements)

References 75 publications

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“…In the present study, a simple but efficient method was selected, which was previously described and calibrated for natural soil samples 17, 43 . While, as reported by Frossard et al .…”

Section: Discussionmentioning

confidence: 99%

Soil bacterial quantification approaches coupling with relative abundances reflecting the changes of taxa

Zhang

et al. 2017

Sci Rep

126

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Understanding the abundance change of certain bacterial taxa is quite important for the study of soil microbiology. However, the observed differences of relative abundances by high-throughput techniques may not accurately reflect those of the actual taxon abundances. This study investigated whether soil microbial abundances coupling with microbial quantities can be more informative in describing the microbial population distribution under different locations. We analyzed relative abundances of the major species in soil microbial communities from Beijing and Tibet grasslands by using 16 S rRNA high-throughput sequencing technique, and quantified the absolute bacterial cell numbers directly or indirectly by multiple culture-independent measurements, including adenosine tri-phosphate (ATP), flow cytometry (FCM), quantitative real-time PCR (qPCR), phospholipid fatty acids (PLFA) and microbial biomass Carbon (MBC). By comparison of the relative abundance and the estimated absolute abundances (EAA) of the major components in soil microbial communities, several dominant phyla, including Actinobacteria, Bacteroidetes, Verrucomicrobia, Chloroflexi, Gemmatimonates and Planctomycetes, showed significantly different trends. These results indicated that the change in EAA might be more informative in describing the dynamics of a population in a community. Further studies of soil microbes should combine the quantification and relative abundances of the microbial communities for the comparisons among various locations.

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“…In the present study, a simple but efficient method was selected, which was previously described and calibrated for natural soil samples 17, 43 . While, as reported by Frossard et al .…”

Section: Discussionmentioning

confidence: 99%

Soil bacterial quantification approaches coupling with relative abundances reflecting the changes of taxa

Zhang

et al. 2017

Sci Rep

126

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“…Over the last decades, flow cytometry has become a valuable tool within the field of environmental microbiology 27 28 . This cellular technique has been used to quantify bacterial numbers throughout the entire range of growth phases under a large variety of physiological conditions and for microbes living in a wide diversity of habitats: soil 29 , water 30 or food 31 .…”

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confidence: 99%

Direct flow cytometry measurements reveal a fine-tuning of symbiotic cell dynamics according to the host developmental needs in aphid symbiosis

Simonet

Duport

Gaget

et al. 2016

Sci Rep

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Endosymbiotic associations constitute a driving force in the ecological and evolutionary diversification of metazoan organisms. Little is known about whether and how symbiotic cells are coordinated according to host physiology. Here, we use the nutritional symbiosis between the insect pest, Acyrthosiphon pisum, and its obligate symbiont, Buchnera aphidicola, as a model system. We have developed a novel approach for unculturable bacteria, based on flow cytometry, and used this method to estimate the absolute numbers of symbionts at key stages of aphid life. The endosymbiont population increases exponentially throughout nymphal development, showing a growing rate which has never been characterized by indirect molecular techniques. Using histology and imaging techniques, we have shown that the endosymbiont-bearing cells (bacteriocytes) increase significantly in number and size during the nymphal development, and clustering in the insect abdomen. Once adulthood is reached and the laying period has begun, the dynamics of symbiont and host cells is reversed: the number of endosymbionts decreases progressively and the bacteriocyte structure degenerates during insect aging. In summary, these results show a coordination of the cellular dynamics between bacteriocytes and primary symbionts and reveal a fine-tuning of aphid symbiotic cells to the nutritional demand imposed by the host physiology throughout development.

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“…5 g paddy soil was diluted to 50 ml with 0.9% NaCl solution and vortexed for 5 min after which samples were centrifuged at 130 g for 5 min. The supernatant was filtered (300-mesh filter) to remove particulate matter (Bressan et al 2015). This suspension was diluted to 10 6 cells per ml and 0.5 ml was added to a flow tube after which 5 μl of both TO and PI solutions was added (Fig.…”

Section: Ichip Manufacture and Single Cell Sortingmentioning

confidence: 99%

FACS-iChip: a high-efficiency iChip system for microbial ‘dark matter’ mining

Liu

Xue

Yi-ling

et al. 2020

Mar Life Sci Technol

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The isolation chip method (iChip) provides a novel approach for culturing previously uncultivable microorganisms; this method is currently limited by the user being unable to ensure single-cell loading within individual wells. To address this limitation, we integrated flow cytometry-based fluorescence-activated cell sorting with a modified iChip (FACS-iChip) to effectively mine microbial dark matter in soils. This method was used for paddy soils with the aim of mining uncultivable microorganisms and making preliminary comparisons between the cultured microorganisms and the bulk soil via 16S rRNA gene sequencing. Results showed that the FACS-iChip achieved a culture recovery rate of almost 40% and a culture retrieval rate of 25%. Although nearly 500 strains were cultured from 19 genera with 8 FACS-iChip plates, only six genera could be identified via 16S rRNA gene amplification. This result suggests that the FACS-iChip is capable of detecting strains in the currently dead spaces of PCR-based sequencing technology. We, therefore, conclude that the FACS-iChip system provides a highly efficient and readily available approach for microbial 'dark matter' mining. Keywords FACS-iChip • Microbial 'dark matter' • In situ cultivation • Single cell sorting SPECIAL TOPIC: Cultivation of the uncultured microorganisms. Haoze Liu, Ran Xue and Yiling Wang contributed equally. Edited by Chengchao Chen.

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A rapid flow cytometry method to assess bacterial abundance in agricultural soil

Cited by 41 publications

References 75 publications

Soil bacterial quantification approaches coupling with relative abundances reflecting the changes of taxa

Soil bacterial quantification approaches coupling with relative abundances reflecting the changes of taxa

Direct flow cytometry measurements reveal a fine-tuning of symbiotic cell dynamics according to the host developmental needs in aphid symbiosis

FACS-iChip: a high-efficiency iChip system for microbial ‘dark matter’ mining

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