A methodological review on the characterization of microalgal biofilm and its extracellular polymeric substances

Abstract

Biofilm secreted by microalgae are extracellular polymeric substances (EPSs) composed mainly of polysaccharides, proteins, nucleic acids and lipids. These EPSs immobilize the cells and stabilize biofilm, mediating adhesion towards solid surfaces.The EPSs valorization through industrial exploitations and scientific works is becoming more popular, but the bottleneck of such studies is the lack of consensus among researchers on the selection of detection techniques to be used, especially for novice researchers. It is a daunting task for any inexperienced researcher when they fail to identify the right tools needed for microalgal biofilm studies. In this review, a wellrefined analysis protocol about microalgal biofilm and EPSs were prepared including its extraction and characterization. Pros and cons of various detection techniques were addressed and cutting-edge methods to study biofilm EPSs were highlighted.Future perspectives were also presented at the end of this review to bridge research gaps in studying biofilm adhesion via EPSs production. Ultimately, this review aims to assist novice researchers in making the right choices in their research studies on microalgal biofilms in accordance to the available technologies and needs.

Polysaccharides’ Structures and Functions in Biofilm Architecture of Antimicrobial-Resistant (AMR) Pathogens

“…As a result of their specificity and the need of few sample manipulations, fluorescent-labelled lectins have been suggested as effective CLSM in situ probes and can be used not only for visualization but also for the semiquantitative analysis. The usefulness of various lectins has been widely demonstrated, starting from the most frequently used concanavalin A (ConA) and wheat germ agglutinin (WGA) [ 124 ]. Lectin selection is the key point of exopolysaccharides’ detection; however, due to their narrow specificity, the choice could be challenging.…”

“…To evaluate the polysaccharide composition, the glycosidic bonds must be previously hydrolyzed by enzymes or acid treatment. Thereafter, monosaccharide composition and quantity can be determined by high-performance liquid chromatography (HPLC) [ 124 ] or high-performance ion exchange chromatography with pulse amperometric detection (HPAEC-PAD) [ 133 ]. In order to have information not only on the composition but also on glycosyl linkages, a sample permethylated by the Ciukanu and Kerek method [ 134 ], hydrolyzed with trifluoroacetic acid, reduced with NaBH 4 and acetylated can be analyzed by gas chromatography coupled with mass spectrometry (GC-MS) [ 133 , 135 ].…”

Fourier Transform Infrared (FTIR) Spectroscopic Study of Biofilms Formed by the Rhizobacterium Azospirillum baldaniorum Sp245: Aspects of Methodology and Matrix Composition

“…The diverse field of microbiology has been progressively developing over the last decades, largely owing to the steadily growing applications of a range of modern molecular spectroscopy techniques that provide molecular-level information on complicated microbiological objects and allow for successfully solving various bioanalytical problems, often in situ or in vivo (see, e.g., [ 1 , 2 , 3 , 4 , 5 , 6 , 7 , 8 , 9 , 10 , 11 , 12 , 13 , 14 , 15 , 16 , 17 ]). We would like to emphasize the informativity of very relevant vibrational spectroscopic techniques, including various modifications of infrared (conventional absorption [ 2 , 7 , 11 , 16 ], diffuse reflectance [ 3 , 6 ], attenuated total reflectance (ATR) [ 5 , 17 ] and surface-enhanced absorption [ 15 ] modes; 2D infrared [ 8 ]) and Raman [ 1 , 2 , 13 ] spectroscopies, as well as their combinations [ 4 , 14 ]; vibrational circular dichroism [ 12 ], optical activity [ 9 ], etc. Of special notice are microbiological applications of Mössbauer spectroscopy, in both transmission and emission variants [ 2 , 10 ].…”

“…Among these techniques, Fourier transform infrared (FTIR) spectroscopy, which has already become a routine tool in material science, yet could not be regarded as routine in the life sciences [ 4 , 6 , 7 ], and particularly in microbiology [ 3 , 5 , 8 , 13 , 14 , 15 , 17 , 18 , 19 ]. This situation can in part be explained by the intrinsically sophisticated compositions and structures of diverse microbiological objects, making the processes of sample treatment or preparation for spectroscopic studies, together with treatment of the spectroscopic data and interpreting the results, a challenging task [ 19 ].…”

“…Moreover, the overall cell composition can also rapidly alter depending on the growth phase and/or in response to the changing environments. This complexity is probably the main reason that, despite a great number of studies involving FTIR spectroscopy in microbiology reported so far (see, e.g., [ 3 , 5 , 8 , 14 , 15 , 17 , 18 , 19 ]), the available standardised protocols for FTIR spectroscopic microbiological analyses (including sample preparation) are insufficient, not yet fully generalised and cannot cover all possible cases [ 19 , 20 , 21 , 22 ]. Thus, in the review chapter by Ojeda and Dittrich [ 19 ], a general overview was presented covering the different sample preparation protocols for infrared spectroscopic analysis of bacterial cells, the basic principles of the technique, procedures for calculating vibrational frequencies based on simple harmonic motions, as well as the advantages and disadvantages of FTIR spectroscopy for the analysis of microorganisms.…”