Individual Microcystis colonies harbour distinct bacterial communities that differ by Microcystis oligotype and with time.

Interactions between bacteria and phytoplankton in the phycosphere have impacts at the scale of whole ecosystems, including the development of harmful algal blooms. The cyanobacterium Microcystis causes toxic blooms that threaten freshwater ecosystems and human health globally. Microcystis grows in colonies that harbour dense assemblages of other bacteria, yet the taxonomic composition of these phycosphere communities and the nature of their interactions with Microcystis are not well characterized. To identify the taxa and compositional variance within Microcystis phycosphere communities, we performed 16S rRNA V4 region amplicon sequencing on individual Microcystis colonies collected biweekly via high-throughput droplet encapsulation during a western Lake Erie cyanobacterial bloom. The Microcystis phycosphere communities were distinct from microbial communities in whole water and bulk phytoplankton seston in western Lake Erie but lacked 'core' taxa found across all colonies. However, dissimilarity in phycosphere community composition correlated with sampling date and the Microcystis 16S rRNA oligotype. Several taxa in the phycosphere were specific to and conserved with Microcystis of a single oligotype or sampling date. Together, this suggests that physiological differences between Microcystis strains, temporal changes in strain phenotypes, and the composition of seeding communities may impact community composition of the Microcystis phycosphere.

Anaerobic Disposal of Arsenic-Bearing Wastes Results in Low Microbially Mediated Arsenic Volatilization.

The removal of arsenic from drinking water sources produces arsenic-bearing wastes, which are disposed of in a variety of ways. Several disposal options involve anaerobic environments, including mixing arsenic waste with cow dung, landfills, anaerobic digesters, and pond sediments. Though poorly understood, the production of gaseous arsenic species in these environments can be a primary goal (cow dung mixing) or an unintended consequence (anaerobic digesters). Once formed, these gaseous arsenic species are readily diluted in the atmosphere. Arsenic volatilization can be mediated by the enzyme arsenite S-adenosylmethionine methyltransferase (ArsM) or through the enzymes involved in methanogenesis. In this study, methanogenic mesocosms with arsenic-bearing ferric iron waste from an electrocoagulation drinking water treatment system were used to evaluate the role of methanogenesis in arsenic volatilization using methanogen inhibitors. Arsenic volatilization was highest in methanogenic mesocosms, but represented <0.02% of the total arsenic added. 16S rRNA cDNA sequencing, qPCR of mcrA transcripts, and functional gene array-based analysis of arsM expression, revealed that arsenic volatilization correlated with methanogenic activity. Aqueous arsenic concentrations increased in all mesocosms, indicating that unintended contamination may result from disposal in anaerobic environments. This highlights that more research is needed before recommending anaerobic disposal intended to promote arsenic volatilization.

The effect of droplet size on syntrophic dynamics in droplet-enabled microbial co-cultivation.

Co-cultivation in microfluidic droplets has emerged as a versatile tool for the study of natural and synthetic microbial communities. In particular, the identification and characterization of syntrophic interactions in these communities is attracting increasing interest due to their critical importance for the functioning of environmental and host-associated communities as well as new biotechnological applications. However, one critical parameter in droplet-enabled co-cultivation that has evaded appropriate evaluation is the droplet size. Given the same number of initial cells, a larger droplet size can increase the length scale secreted metabolites must diffuse as well as dilute the initial concentration of cells and exchanged metabolites, impacting the community dynamics. To evaluate the effect of droplet size on a spectrum of syntrophic interactions, we cultivated a synthetic model system consisting of two E. coli auxotrophs, whose interactions could be modulated through supplementation of related amino acids in the medium. Our results demonstrate that the droplet size impacts substantially numerous aspects of the growth of a cross-feeding bi-culture, particularly the growth capacity, maximum specific growth rate, and lag time, depending on the degree of the interaction. This work heavily suggests that one droplet size does not fit all types of interactions; this parameter should be carefully evaluated and chosen in experimental studies that aim to utilize droplet-enabled co-cultivation to characterize or elucidate microbial interactions.

Co-cultivation of microbial sub-communities in microfluidic droplets facilitates high-resolution genomic dissection of microbial 'dark matter'.

While the 'unculturable' majority of the bacterial world is accessible with culture-independent tools, the inability to study these bacteria using culture-dependent approaches has severely limited our understanding of their ecological roles and interactions. To circumvent cultivation barriers, we utilize microfluidic droplets as localized, nanoliter-size bioreactors to co-cultivate subsets of microbial communities. This co-localization can support ecological interactions between a reduced number of encapsulated cells. We demonstrated the utility of this approach in the encapsulation and co-cultivation of droplet sub-communities from a fecal sample collected from a healthy human subject. With the whole genome amplification and metagenomic shotgun sequencing of co-cultivated sub-communities from 22 droplets, we observed that this approach provides accessibility to uncharacterized gut commensals for study. The recovery of metagenome-assembled genomes from one droplet sub-community demonstrated the capability to dissect the sub-communities with high-genomic resolution. In particular, genomic characterization of one novel member of the family Neisseriaceae revealed implications regarding its participation in fatty acid degradation and production of atherogenic intermediates in the human gut. The demonstrated genomic resolution and accessibility to the microbial 'dark matter' with this methodology can be applied to study the interactions of rare or previously uncultivated members of microbial communities.