Environmental Sampling of Photosynthetic Microbes and Their Viruses: From Field to Lab

Sobroney Heng; Ruben Michael Ceballos

doi:10.3791/68379

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Method Article

Environmental Sampling of Photosynthetic Microbes and Their Viruses: From Field to Lab

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DOI:

10.3791/68379

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July 3rd, 2025

Sobroney Heng^*¹ , Ruben Michael Ceballos^*¹^,²

¹Department of Molecular and Cell Biology, University of California, Merced, ²Quantitative Systems Biology Program, University of California, Merced

^* These authors contributed equally

Summary

Here, we describe methods of sampling from the euphotic zones of marine and freshwater ecosystems to isolate cyanophages (and their hosts) and post-sampling processes that enrich single genotypes and characterize virus-host interaction dynamics.

Abstract

Environmental sampling of photosynthetic microorganisms and their viruses plays a critical role in understanding contemporary marine and freshwater biodiversity and ecosystem dynamics as well as the impacts of climate change-related factors (e.g., rising temperatures and acidification) on evolutionary trajectories of species and community composition. Unfortunately, the diversity of the virosphere does not support a single universal sampling and experimental workflow. Indeed, each virus system has unique features, which require modifications to standard protocols in virology to accomplish research goals. Although virus discovery and characterization require approaches that are specific to the target system, for all viruses, the research aims are similar: isolate the virus; determine host range; confirm productive infection; and characterize the virus, the host, and virus-host dynamics. Robust descriptions of virus-host systems consist minimally of elucidating morphology, physiology, biochemistry, and omics profiles. Further information may be obtained by manipulating the system by changing factors such as multiplicity of infection, temperature, pH, host-switch, directed evolution, or applying drugs to observe virus-host system response. Our laboratory studies viruses across domains of life (Archaea, Bacteria, and Eukarya). In this report, we detail methods for sampling photosynthetic microbes from the euphotic zone of freshwater and marine environments with focus on isolating bacteriophage (i.e., cyanophage) of cyanobacteria. Cyanobacteria are keystone species critical to primary production and nutrient cycling in these aquatic ecosystems. The described workflow extends from sampling waters at different depths to characterizing virus-host system features using liquid and solid media culture, advanced molecular/genetic methods, and analytical approaches. The methods described are adaptable to bacteriophage and virus discovery in virus-host systems across domains of life.

Introduction

Characterizing infection dynamics and evolutionary relationships of cyanophages and their hosts is key to understanding the current and future states of marine and freshwater ecosystems. Advanced high-throughput sequencing techniques and modern analytical methods allow rapid sequencing of cyanophage (and host) genomes to study virus-host relationships¹^,². However, workflows begin with environmental sampling to isolate and identify viruses and their hosts³^,⁴^,⁵. Due to the diversity of the virosphere, there is no single....