Confocal Laser Scanning Microscopy Image Analysis for Cyanobacterial Biomass Determined at Microscale Level in Different Microbial Mats

Solé, Antonio; Diestra, Elia; Esteve, Isabel

doi:10.1007/s00248-008-9463-y

Cited by 18 publications

(15 citation statements)

References 31 publications

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“…According to Solé et al . () for the individual of biomass determination of C. chilense , the structures were screened in each original stack. In each z‐stack, a structure was initially selected and then erased from the stack.…”

Section: Methodsmentioning

confidence: 99%

Cyanidium chilense (Cyanidiophyceae, Rhodophyta) from tuff rocks of the archeological site of Cuma, Italy

Ciniglia

Cennamo

Natale

et al. 2019

Phycological Research

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SUMMARY Phlegrean Fields is a large volcanic area situated southwest of Naples (Italy), including both cave and thermoacidic habitats. These extreme environments host the genus Cyanidium; the species C. chilense represents a common phototrophic microorganism living in anthropogenic caves. With a view to provide a comprehensive characterization for a correct taxonomic classification, morpho‐ultrastructural investigations of C. chilense from Sybil's cave (Phlegren Fields) was herein carried out and compared with the thermoacidophilic C. caldarium. The biofilm was also analyzed to define the role of C. chilense in the establishment of a biofilm within cave environments. Despite the peculiar ecological and molecular divergences, C. chilense and C. caldarium shared all the main diacritic features, suggesting morphological convergence within the genus; cytological identity was found among C. chilense strains geographically distant and adapted to different substrates, such as the porous yellow tuff of Sybil cave, and calcyte, magnesite and basaltic rocks from other caves. C. chilense is generally dominant in all biofilms, developing monospecific islets, developing both superficially or between fungal hyphae and coccoid cyanobacteria. Extracellular polymeric substances (EPS) were recorded in C. chilense biofilms from Sybil cave, confirming the role of EPS in facilitating cells adhesion to the surface, creating a cohesive network of interconnecting biofilm cells.

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Section: Methodsmentioning

confidence: 99%

Cyanidium chilense (Cyanidiophyceae, Rhodophyta) from tuff rocks of the archeological site of Cuma, Italy

Ciniglia

Cennamo

Natale

et al. 2019

Phycological Research

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“…Thus, it is necessary to improve our knowledge regarding complex BSC interactions. Fluorescence microscopy has been applied repeatedly to cryptogamic organisms since photoautotrophic organisms emit naturally auto fluorescence (Schallenberg et al, 1989;Kuwae and Hosokawa 1999;Solé et al, 2009;Raanan et al, 2016). The confocal laser scanning microscopy (CLSM) technique applied in this work gives novel insights into the complex architecture of BSCs, avoiding the need to either manipulate or stain 15 the samples.…”

Section: Introductionmentioning

confidence: 99%

“…Moreover, it allows accurate and non-destructive optical investigations of BSC cross sections that generate high resolution images where out-of-focus is eliminated. As a highly comparable approach, it is possible for the first time to calculate C values and partitioning patterns of green algae and cyanobacteria within intact BSCs by applying the CLSM image analysis (CLSM-IA) procedure developed by the group of Solé (2009). Additionally, these results were set in relation to organic C values obtained by loss on ignition to highlight the CO2 sink character of the first cm³ of these cold soils dominated 20 by BSCs.…”

Section: Introductionmentioning

confidence: 99%

Uncovering biological soil crusts: Carbon content and structure of intact Arctic, Antarctic and alpine biological soil crusts

Jung¹,

Briegel-Williams²,

Simon³

et al. 2017

Preprint

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Abstract. Arctic, Antarctic and alpine biological soil crusts (BSCs) are formed by adhesion of soil particles to exopolysaccharides (EPS), excreted by cyanobacterial and green algal communities, the pioneers and main producers in these habitats. These BSCs provide and influence many ecosystem services such as soil erodibility, soil formation and Nitrogen-(N) as well as carbon-(C) cycles. In cold environments degradation rates are low and BSCs increase continuously soil organic C, whereby these soils are considered as CO2 sinks. This work provides a novel, non-destructive and highly comparable method 15 to investigate intact BSCs with a focus on cyanobacteria and green algae and their contribution to soil organic C. A new terminology arose, based on confocal laser scanning microscopy (CLSM) 2D biomaps, dividing BSCs into a photosynthetic active layer (PAL), made of active photoautotrophic organisms and a photosynthetic inactive layer (PIL), harbouring remnants of cyanobacteria and green algae glued together by their remaining EPS. By the application of CLSM image analysis (CLSM-IA) to 3D biomaps, C coming from photosynthetic active organisms could be visualized as depth profiles with C peaks at 0.5 20 to 2 mm depth. Additionally, the CO2 sink character of these cold soil habitats dominated by BSCs could be highlighted, demonstrating that the first cm³ of soil is made of between 7 and 17 % total organic carbon, identified by loss on ignition.

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“…It also offers the possibility of non-invasive optical sectioning by subtracting out-of-focus planes of the image [9,14]. By exploiting natural pigment fluorescence emitted by cyanobacteria (chlorophyll-a and phycobilins) with an emission wavelength of 665 nm, CLSM can be used for their detection in any sample [17]. In spite of the potential of this technique to study cyanobacteria, there are some limitations to its application to the study of plant-microbe interactions.…”

Section: Introductionmentioning

confidence: 99%

DTAF: an efficient probe to study cyanobacterial-plant interaction using confocal laser scanning microscopy (CLSM)

Ahmed

Stal

Hasnain

2010

J Ind Microbiol Biotechnol

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A variety of microscopic techniques have been utilized to study cyanobacterial associations with plant roots, but confocal laser scanning microscopy (CLSM) is the least used due to the unavailability of a suitable fluorescent dye. Commonly used lectins have problems with their binding ability with root cells and their visualization under CLSM. DTAF (5-(4,6-dichlorotriazinyl) aminofluorescein) is a fluorescent dye that has been widely used for staining various biological samples for fluorescent microscopy. It reacts with polysaccharides and peptides at ordinary conditions. The possible application and efficiency of DTAF for CLSM studies were examined in various aspects of cyanobacterial-plant interactions. Seedlings of Pisum sativum, Vigna rediata and Triticum aestivum were co-cultivated and stained with DTAF as a fluorochrome. Extracellular and intracellular interactions of cyanobacteria and the plant root surface were observed by CLSM. Results were compared with staining by other commonly used lectins. Advantages of the use of DTAF over other stains are its penetration into root tissues and binding with polysaccharides, mainly the cellulose. The staining was smooth, which clearly showed minute details on the cell of surface and root hairs with higher resolution. The emission wavelength for DTAF is 517 nm, which is highly advantageous as cyanobacteria have auto-fluorescence at 665 nm, and both can be simultaneously used in CLSM by visualizing in different channels. This worked efficiently with all three plants used and with filamentous and unicellular cyanobacterial strains. Cyanobacterial presence was not only clearly observed on the root surface, but also inside the root tissue and epidermal cells. The easy protocol and absence of tissue processing make DTAF a useful probe for studies of cyanobacterial associations with plant roots by CLSM.

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Confocal Laser Scanning Microscopy Image Analysis for Cyanobacterial Biomass Determined at Microscale Level in Different Microbial Mats

Cited by 18 publications

References 31 publications

Cyanidium chilense (Cyanidiophyceae, Rhodophyta) from tuff rocks of the archeological site of Cuma, Italy

Cyanidium chilense (Cyanidiophyceae, Rhodophyta) from tuff rocks of the archeological site of Cuma, Italy

Uncovering biological soil crusts: Carbon content and structure of intact Arctic, Antarctic and alpine biological soil crusts

DTAF: an efficient probe to study cyanobacterial-plant interaction using confocal laser scanning microscopy (CLSM)

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