A protocol for the simultaneous identification of chitin-containing particles and their associated bacteria
Florencia Biancalana
Germán A. Kopprio
Rubén J. Lara
Cecilia Alonso
Abstract
Chitin is the second most abundant polymer on Earth, playing a crucial role in the biogeochemical cycles. A core issue for studying its processing in aquatic systems is the identification and enumeration of chitin-containing particles and organisms, ideally in a manner that can be directly linked to bulk chitin quantification. The aim of this study was the development of such a technique. We successfully combined the methodology of bulk chitin determination using wheat germ agglutinin (FITC-WGA) for staining chitin-containing particles and organisms along with CARD-FISH staining of either chitin-containing eukaryotic cells or bacteria associated with them. Environmental chitin staining was successfully applied to natural water samples. Fungal hyphae, diatoms, and dinoflagellates, sestonic aggregates and chitin-containing structures derived from metazoa were observed. Also, hybridized bacteria attached to chitinaceous debris were clearly visualized. Finally, as proof of principle, cultured yeast cells were simultaneously-targeted by FITC-WGA and the EUK516 probe without exhibiting any interference between both stains. The presented approach appears as a powerful tool to evaluate the contribution of different size classes and organisms to chitin production and consumption, opening the possibility for application of single-cell approaches targeting the ecophysiology of chitin transformations in aquatic systems.
Biancalana F., Kopprio G.A., Lara R.J. and Alonso C. (2017) A protocol for the simultaneous identification of chitin-containing particles and their associated bacteria. 40(5): 314-320. 10.1016/j.syapm.2017.05.004
@article{Biancalana2017,
Title = {A protocol for the simultaneous identification of chitin-containing particles and their associated bacteria},
Author = {Biancalana, Florencia and Kopprio, Germán A. and Lara, Rubén J. and Alonso, Cecilia},
Editor = {},
Year = {2017},
Pages = {314-320},
Volume = {40},
Doi = {10.1016/j.syapm.2017.05.004},
Abstract = {Chitin is the second most abundant polymer on Earth, playing a crucial role in the biogeochemical cycles. A core issue for studying its processing in aquatic systems is the identification and enumeration of chitin-containing particles and organisms, ideally in a manner that can be directly linked to bulk chitin quantification. The aim of this study was the development of such a technique. We successfully combined the methodology of bulk chitin determination using wheat germ agglutinin (FITC-WGA) for staining chitin-containing particles and organisms along with CARD-FISH staining of either chitin-containing eukaryotic cells or bacteria associated with them. Environmental chitin staining was successfully applied to natural water samples. Fungal hyphae, diatoms, and dinoflagellates, sestonic aggregates and chitin-containing structures derived from metazoa were observed. Also, hybridized bacteria attached to chitinaceous debris were clearly visualized. Finally, as proof of principle, cultured yeast cells were simultaneously-targeted by FITC-WGA and the EUK516 probe without exhibiting any interference between both stains. The presented approach appears as a powerful tool to evaluate the contribution of different size classes and organisms to chitin production and consumption, opening the possibility for application of single-cell approaches targeting the ecophysiology of chitin transformations in aquatic systems.},
}
TY - JOUR
AU - Biancalana, Florencia
AU - Kopprio, Germán A.
AU - Lara, Rubén J.
AU - Alonso, Cecilia
TI - A protocol for the simultaneous identification of chitin-containing particles and their associated bacteria
PY - 2017
SP - 314-320
VL - 40
DO - 10.1016/j.syapm.2017.05.004
AB - Chitin is the second most abundant polymer on Earth, playing a crucial role in the biogeochemical cycles. A core issue for studying its processing in aquatic systems is the identification and enumeration of chitin-containing particles and organisms, ideally in a manner that can be directly linked to bulk chitin quantification. The aim of this study was the development of such a technique. We successfully combined the methodology of bulk chitin determination using wheat germ agglutinin (FITC-WGA) for staining chitin-containing particles and organisms along with CARD-FISH staining of either chitin-containing eukaryotic cells or bacteria associated with them. Environmental chitin staining was successfully applied to natural water samples. Fungal hyphae, diatoms, and dinoflagellates, sestonic aggregates and chitin-containing structures derived from metazoa were observed. Also, hybridized bacteria attached to chitinaceous debris were clearly visualized. Finally, as proof of principle, cultured yeast cells were simultaneously-targeted by FITC-WGA and the EUK516 probe without exhibiting any interference between both stains. The presented approach appears as a powerful tool to evaluate the contribution of different size classes and organisms to chitin production and consumption, opening the possibility for application of single-cell approaches targeting the ecophysiology of chitin transformations in aquatic systems.
ER -
