Viruses affect picocyanobacterial abundance and biogeography in the North Pacific Ocean.

The photosynthetic picocyanobacteria Prochlorococcus and Synechococcus are models for dissecting how ecological niches are defined by environmental conditions, but how interactions with bacteriophages affect picocyanobacterial biogeography in open ocean biomes has rarely been assessed. We applied single-virus and single-cell infection approaches to quantify cyanophage abundance and infected picocyanobacteria in 87 surface water samples from five transects that traversed approximately 2,200 km in the North Pacific Ocean on three cruises, with a duration of 2-4 weeks, between 2015 and 2017. We detected a 550-km-wide hotspot of cyanophages and virus-infected picocyanobacteria in the transition zone between the North Pacific Subtropical and Subpolar gyres that was present in each transect. Notably, the hotspot occurred at a consistent temperature and displayed distinct cyanophage-lineage composition on all transects. On two of these transects, the levels of infection in the hotspot were estimated to be sufficient to substantially limit the geographical range of Prochlorococcus. Coincident with the detection of high levels of virally infected picocyanobacteria, we measured an increase of 10-100-fold in the Synechococcus populations in samples that are usually dominated by Prochlorococcus. We developed a multiple regression model of cyanophages, temperature and chlorophyll concentrations that inferred that the hotspot extended across the North Pacific Ocean, creating a biological boundary between gyres, with the potential to release organic matter comparable to that of the sevenfold-larger North Pacific Subtropical Gyre. Our results highlight the probable impact of viruses on large-scale phytoplankton biogeography and biogeochemistry in distinct regions of the oceans.

A single-cell polony method reveals low levels of infected Prochlorococcus in oligotrophic waters despite high cyanophage abundances.

Long-term stability of picocyanobacteria in the open oceans is maintained by a balance between synchronous division and death on daily timescales. Viruses are considered a major source of microbial mortality, however, current methods to measure infection have significant methodological limitations. Here we describe a method that pairs flow-cytometric sorting with a PCR-based polony technique to simultaneously screen thousands of taxonomically resolved individual cells for intracellular virus DNA, enabling sensitive, high-throughput, and direct quantification of infection by different virus lineages. Under controlled conditions with picocyanobacteria-cyanophage models, the method detected infection throughout the lytic cycle and discriminated between varying infection levels. In North Pacific subtropical surface waters, the method revealed that only a small percentage of Prochlorococcus (0.35-1.6%) were infected, predominantly by T4-like cyanophages, and that infection oscillated 2-fold in phase with the diel cycle. This corresponds to 0.35-4.8% of Prochlorococcus mortality daily. Cyanophages were 2-4-fold more abundant than Prochlorococcus, indicating that most encounters did not result in infection and suggesting infection is mitigated via host resistance, reduced phage infectivity and inefficient adsorption. This method will enable quantification of infection for key microbial taxa across oceanic regimes and will help determine the extent that viruses shape microbial communities and ecosystem level processes.

Quantification of T4-Like and T7-Like Cyanophages Using the Polony Method Show They Are Significant Members of the Virioplankton in the North Pacific Subtropical Gyre.

The North Pacific Subtropical Gyre (NPSG) is one of the largest biomes on Earth, with the cyanobacterium Prochlorococcus being the most abundant primary producer year-round. Viruses that infect cyanobacteria (cyanophages) influence cyanobacterial mortality, diversity and evolution. Two major cyanophage families are the T4-like cyanomyoviruses and T7-like cyanopodoviruses, yet their abundances and distribution patterns remain unknown due to difficulty in quantifying their populations. To address this limitation, we previously adapted the polony method (for PCR colony) to quantify T7-like cyanophages and applied it to spring populations in the Red Sea. Here, we further adapted the method for the quantification of T4-like cyanophages and analyzed the abundances of T4-like and T7-like cyanophage populations in the photic zone of the NPSG in summer 2015 and spring 2016. Combined, the peak abundances of these two cyanophage families reached 2.8 × 106 and 1.1 × 106 cyanophages ⋅ ml-1 in the summer and spring, respectively. They constituted between 3 and 16% of total virus-like particles (VLPs), comprising a substantial component of the virioplankton in the NPSG. While both cyanophage families were highly abundant, the T4-like cyanophages were generally 1.3-4.4 fold more so. In summer, cyanophages had similar and reproducible distribution patterns with depth. Abundances were relatively low in the upper mixed layer and increased to form a pronounced subsurface peak at 100 m (1.9 × 106 and 9.1 × 105 phages ⋅ ml-1 for the T4-like and T7-like cyanophages, respectively), coincident with the maximum in Prochlorococcus populations. Less vertical structure in cyanophage abundances was apparent in the spring profile, despite a subsurface peak in Prochlorococcus numbers. In the summer upper mixed layer, cyanophages constituted a smaller proportion of VLPs than below it and cyanophage to cyanobacteria ratios were considerably lower (1.3-2.8) than those of VLPs to bacteria (8.1-21.2). Differences in abundances between the two families and their contribution to VLPs with depth suggest differences in cyanophage production and/or decay processes relative to other members of the virioplankton in the upper mixed layer. These findings highlight the importance of quantifying distinct populations within the virioplankton to gain accurate understanding of their distribution patterns.

Single cell genomes of Prochlorococcus, Synechococcus, and sympatric microbes from diverse marine environments.

Prochlorococcus and Synechococcus are the dominant primary producers in marine ecosystems and perform a significant fraction of ocean carbon fixation. These cyanobacteria interact with a diverse microbial community that coexists with them. Comparative genomics of cultivated isolates has helped address questions regarding patterns of evolution and diversity among microbes, but the fraction that can be cultivated is miniscule compared to the diversity in the wild. To further probe the diversity of these groups and extend the utility of reference sequence databases, we report a data set of single cell genomes for 489 Prochlorococcus, 50 Synechococcus, 9 extracellular virus particles, and 190 additional microorganisms from a diverse range of bacterial, archaeal, and viral groups. Many of these uncultivated single cell genomes are derived from samples obtained on GEOTRACES cruises and at well-studied oceanographic stations, each with extensive suites of physical, chemical, and biological measurements. The genomic data reported here greatly increases the number of available Prochlorococcus genomes and will facilitate studies on evolutionary biology, microbial ecology, and biological oceanography.