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Extracellular enzyme activity is a well-established parameter for evaluating microbial biogeochemical roles in marine ecosystems. The presence and activity of extracellular enzymes in seawater provide insights into the quality and quantity of organic matter being processed by the present microorganisms. A key challenge in our understanding of these processes is to decode the extracellular enzyme repertoire and activities of natural communities at the single-cell level. Current measurements are carried out on bulk or size-fractionated samples capturing activities of mixed populations. This approach – even with size-fractionation – cannot be used to trace enzymes back to their producers, nor distinguish the active microbial members, leading to a disconnect between measured activities and the producer cells. By targeting extracellular enzymes and resolving their activities at the single-cell level, we can investigate underlying phenotypic heterogeneity among clonal or closely related organisms, characterize enzyme kinetics under varying environmental conditions, and resolve spatio-temporal distribution of individual enzyme producers within natural communities. In this perspective piece, we discuss state-of-the-art technologies in the fields of microfluidic droplets and functional screening of prokaryotic cells for measuring enzyme activity in marine seawater samples, one cell at a time. We further elaborate on how this single-cell approach can be used to address research questions that cannot be answered with current methods, as pertinent to the enzymatic degradation of organic matter by marine microorganisms. 

Prophages are known to encode important virulence factors in the human pathogenVibrio cholerae. However, little is known about the occurrence and composition of prophage-encoded traits in environmental vibrios. A database of 5,674 prophage-like elements constructed from 1,874Vibriogenome sequences, covering sixty-four species, revealed that prophage-like elements encoding possible properties such as virulence and antibiotic resistance are widely distributed among environmental vibrios, including strains classified as non-pathogenic. Moreover, we found that 45% ofVibriospecies harbored a complete prophage-like element belonging to theInoviridaefamily, which encode the zonula occludens toxin (Zot) previously described in theV.cholerae. Interestingly, thesezot-encoding prophages were found in a variety ofVibriostrains covering both clinical and marine isolates, including strains from deep sea hydrothermal vents and deep subseafloor sediments. In addition, the observation that a spacer from the CRISPR locus in the marine fish pathogenV.anguillarumstrain PF7 had 95% sequence identity with azotgene from theInoviridaeprophage found inV.anguillarumstrain PF4, suggests acquired resistance to inoviruses in this species. Altogether, our results contribute to the understanding of the role of prophages as drivers of evolution and virulence in the marineVibriobacteria.

 

Increasing glacial discharge can lower salinity and alter organic matter (OM) supply in fjords, but assessing the biogeochemical effects of enhanced freshwater fluxes requires understanding of microbial interactions with OM across salinity gradients. Here, we examined microbial enzymatic capabilities—in bulk waters (nonsize‐fractionated) and on particles (≥ 1.6μm)—to hydrolyze common OM constituents (peptides, glucose, polysaccharides) along a freshwater–marine continuum within Tyrolerfjord‐Young Sound. Bulk peptidase activities were up to 15‐fold higher in the fjord than in glacial rivers, whereas bulk glucosidase activities in rivers were twofold greater, despite fourfold lower cell counts. Particle‐associated glucosidase activities showed similar trends by salinity, but particle‐associated peptidase activities were up to fivefold higher—or, for several peptidases, only detectable—in the fjord. Bulk polysaccharide hydrolase activities also exhibited freshwater–marine contrasts: xylan hydrolysis rates were fivefold higher in rivers, while chondroitin hydrolysis rates were 30‐fold greater in the fjord. Contrasting enzymatic patterns paralleled variations in bacterial community structure, with most robust compositional shifts in river‐to‐fjord transitions, signifying a taxonomic and genetic basis for functional differences in freshwater and marine waters. However, distinct dissolved organic matter (DOM) pools across the salinity gradient, as well as a positive relationship between several enzymatic activities and DOM compounds, indicate that DOM supply exerts a more proximate control on microbial activities. Thus, differing microbial enzymatic capabilities, community structure, and DOM composition—interwoven with salinity and water mass origins—suggest that increased meltwater may alter OM retention and processing in fjords, changing the pool of OM supplied to coastal Arctic microbial communities.