Microbial Processing and Production of Aquatic Fluorescent Organic Matter in a Model Freshwater System

Abstract

Organic matter (OM) has an essential biogeochemical influence along the hydrological continuum and within aquatic ecosystems. Organic matter derived via microbial processes was investigated within a range of model freshwater samples over a 10-day period. For this, excitation-emission matrix (EEM) fluorescence spectroscopy in combination with parallel factor (PARAFAC) analysis was employed. This research shows the origin and processing of both protein-like and humic-like fluorescence within environmental and synthetic samples over the sampling period. The microbial origin of Peak T fluorescence is demonstrated within both synthetic samples and in environmental samples. Using a range of incubation temperatures provides evidence for the microbial metabolic origin of Peak T fluorescence. From temporally resolved experiments, evidence is provided that Peak T fluorescence is an indication of metabolic activity at the microbial community level and not a proxy for bacterial enumeration. This data also reveals that humic-like fluorescence can be microbially derived in situ and is not solely of terrestrial origin, likely to result from the upregulation of cellular processes prior to cell multiplication. This work provides evidence that freshwater microbes can engineer fluorescent OM, demonstrating that microbial communities not only process, but also transform, fluorescent organic matter.

Laboratory In-Situ Production of Autochthonous and Allochthonous Fluorescent Organic Matter by Freshwater Bacteria

“…Peaks C and C+ are associated with high molecular weight, terrestrially derived, allochthonous organic material [ 15 , 20 , 26 ]. Previous work has shown that microbes can produce Peaks C and C+ in simple matrices [ 38 , 39 , 40 ]. In this study, for the microbial community cultures, Peak C was found to be predominantly associated with extracellular material.…”

“…All fluorescence spectra were blank subtracted using sterile media, corrected for inner filter effects (for both excitation and emission wavelengths) and first and second order Rayleigh Scattering masked (±10 nm at λ ex = λ em and 2λ ex = λ em ) [ 8 , 39 , 40 , 51 ] within the Aqualog ® software (Horiba Ltd., Kyoto, Japan). All fluorescence intensity data were converted and reported in quinine sulphate units (QSU) (1 QSU = 1 µg −1 quinine sulphate) [ 19 , 39 , 40 , 52 , 53 ] via a custom script, written in Python TM (Python Software Foundation, Wilmington, DE, USA).…”

“…A custom script, written in Python TM (Python Software Foundation, Wilmington, DE, USA), was used to create the EEM maps and to undertake the peak picking for specific fluorescence regions of interest [ 39 , 40 ]. The data window of the EEMs was λ ex 240–490 nm, λ em 250–550 nm.…”

“…Although there has been some exploration of the autochthonous origin of microbial FOM outside of marine research, this has been limited to the exploration of single bacterial species [ 38 ] or has used laboratory based-models that employ reference strain microbial inoculums [ 39 , 40 ]. This has demonstrated the ability of microbes to produce a wide range of FOM in-situ, providing an insight into the potential use of Peak T fluorescence as a proxy of microbial activity or the presence of bacteria.…”

Fluorophores in surface freshwaters: importance, likely structures, and possible impacts of climate change

“…43 Proteins and free amino acids can be both consumed and produced by aquatic microorganisms, and are released into the extracellular environment with the lysis of dead cells. The relevant EEM signals are connected to microbial metabolic activity, 44 and they are usually a signature of autochthonous or aquagenic organic matter.…”