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Abstract. Decaying vegetation was determined to be a potentially important source ofatmospheric ice nucleation particles (INPs) in the early 1970s. The bacteriumPseudomonas syringae was the first microorganism with ice nucleationactivity (INA) isolated from decaying leaf litter in 1974. However, the icenucleation characteristics of P. syringae are not compatible withthe characteristics of leaf litter-derived INPs since the latter were foundto be sub-micron in size, while INA of P. syringae depends on muchlarger intact bacterial cells. Here we determined the cumulative icenucleation spectrum and microbial community composition of the historic leaflitter sample 70-S-14 collected in 1970 that conserved INA for 48 years. Themajority of the leaf litter-derived INPs were confirmed to be sub-micron insize and to be sensitive to boiling. Culture-independent microbial communityanalysis only identified Pseudomonas as potential INA.Culture-dependent analysis identified one P. syringae isolate, twoisolates of the bacterial species Pantoea ananatis, and one fungalisolate of Mortierella alpina as having INA among 1170 bacterialcolonies and 277 fungal isolates, respectively. Both Pa. ananatisand M. alpina are organisms that produce heat-sensitive sub-micronINPs. They are thus both likely sources of the INPs present in sample 70-S-14and may represent important terrestrial sources of atmospheric INPs, aconclusion that is in line with other recent results obtained in regard toINPs from soil, precipitation, and the atmosphere. 

Microorganisms are ubiquitous in freshwater aquatic environments, but little is known about their abundance, diversity, and transport. We designed and deployed a remote-operated water-sampling system onboard an unmanned surface vehicle (USV, a remote-controlled boat) to collect and characterize microbes in a freshwater lake in Virginia, USA. The USV collected water samples simultaneously at 5 and 50 cm below the surface of the water at three separate locations over three days in October, 2016. These samples were plated on a non-selective medium (TSA) and on a medium selective for the genusPseudomonas(KBC) to estimate concentrations of culturable bacteria in the lake. Mean concentrations ranged from 134 to 407 CFU/mL for microbes cultured on TSA, and from 2 to 8 CFU/mL for microbes cultured on KBC. There was a significant difference in the concentration of microbes cultured on KBC across three sampling locations in the lake (P= 0.027), suggesting an uneven distribution ofPseudomonasacross the locations sampled. There was also a significant difference in concentrations of microbes cultured on TSA across the three sampling days (P= 0.038), demonstrating daily fluctuations in concentrations of culturable bacteria. There was no significant difference in concentrations of microbes cultured on TSA (P= 0.707) and KBC (P= 0.641) across the two depths sampled, suggesting microorganisms were well-mixed between 5 and 50 cm below the surface of the water. About 1 percent (7/720) of the colonies recovered across all four sampling missions were ice nucleation active (ice+) at temperatures warmer than −10 °C. Our work extends traditional manned observations of aquatic environments to unmanned systems, and highlights the potential for USVs to understand the distribution and diversity of microbes within and above freshwater aquatic environments.