Dataset Information

Pitfalls Associated with Discriminating Mixed-Species Biofilms by Flow Cytometry.

ABSTRACT: Since biofilms are ubiquitous in different settings and act as sources of disease for humans, reliable methods to characterize and quantify these microbial communities are required. Numerous techniques have been employed, but most of them are unidirectional, labor intensive and time consuming. Although flow cytometry (FCM) can be a reliable choice to quickly provide a multiparametric analysis, there are still few applications on biofilms, and even less on the study of inter-kingdom communities. This work aimed to give insights into the application of FCM in order to more comprehensively analyze mixed-species biofilms, formed by different Pseudomonas aeruginosa and Candida albicans strains, before and after exposure to antimicrobials. For comparison purposes, biofilm culturability was also assessed determining colony-forming units. The results showed that some aspects, namely the microbial strain used, the morphological state of the cells and the biofilm matrix, make the accurate analysis of FCM data difficult. These aspects were even more challenging when double-species biofilms were being inspected, as they could engender data misinterpretations. The outcomes draw our attention towards the need to always take into consideration the characteristics of the biofilm samples to be analyzed through FCM, and undoubtedly link to the need for optimization of the processes tailored for each particular case study.

SUBMITTER: Grainha T

PROVIDER: S-EPMC7694060 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Pitfalls Associated with Discriminating Mixed-Species Biofilms by Flow Cytometry.

Grainha Tânia T Magalhães Andreia P AP Melo Luís D R LDR Pereira Maria O MO

Antibiotics (Basel, Switzerland) 20201027 11

Since biofilms are ubiquitous in different settings and act as sources of disease for humans, reliable methods to characterize and quantify these microbial communities are required. Numerous techniques have been employed, but most of them are unidirectional, labor intensive and time consuming. Although flow cytometry (FCM) can be a reliable choice to quickly provide a multiparametric analysis, there are still few applications on biofilms, and even less on the study of inter-kingdom communities. ...[more]

PMID: 33121057

Similar Datasets

Project description:BackgroundOur current understanding of biofilms indicates that these structures are typically composed of many different microbial species. However, the lack of reliable techniques for the discrimination of each population has meant that studies focusing on multi-species biofilms are scarce and typically generate qualitative rather than quantitative data.Methodology/principal findingsWe employ peptide nucleic acid fluorescence in situ hybridization (PNA FISH) methods to quantify and visualize mixed biofilm populations. As a case study, we present the characterization of Salmonella enterica/Listeria monocytogenes/Escherichia coli single, dual and tri-species biofilms in seven different support materials. Ex-situ, we were able to monitor quantitatively the populations of ∼56 mixed species biofilms up to 48 h, regardless of the support material. In situ, a correct quantification remained more elusive, but a qualitative understanding of biofilm structure and composition is clearly possible by confocal laser scanning microscopy (CLSM) at least up to 192 h. Combining the data obtained from PNA FISH/CLSM with data from other established techniques and from calculated microbial parameters, we were able to develop a model for this tri-species biofilm. The higher growth rate and exopolymer production ability of E. coli probably led this microorganism to outcompete the other two [average cell numbers (cells/cm(2)) for 48 h biofilm: E. coli 2,1 × 10(8) (± 2,4 × 10(7)); L. monocytogenes 6,8 × 10(7) (± 9,4 × 10(6)); and S. enterica 1,4 × 10(6) (± 4,1 × 10(5))]. This overgrowth was confirmed by CSLM, with two well-defined layers being easily identified: the top one with E. coli, and the bottom one with mixed regions of L. monocytogenes and S. enterica.SignificanceWhile PNA FISH has been described previously for the qualitative study of biofilm populations, the present investigation demonstrates that it can also be used for the accurate quantification and spatial distribution of species in polymicrobial communities. Thus, it facilitates the understanding of interspecies interactions and how these are affected by changes in the surrounding environment.

Dataset Information

Pitfalls Associated with Discriminating Mixed-Species Biofilms by Flow Cytometry.

Publications

Pitfalls Associated with Discriminating Mixed-Species Biofilms by Flow Cytometry.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets