Ribosome Recapture: A Door to Taxon-specific Microbial Mortality
Xu Zhong1*, Jennifer Wirth1,5*, Curtis Suttle1,2, 3,4
Department of 1Earth, Ocean, and Atmospheric Sciences, 2Microbiology and Immunology, and 3Botany, University of British Columbia, Vancouver, BC, Canada;
4Integrated Microbial Biodiversity Program, Canadian Institute for Advanced Research, Toronto, ON, Canada; 5Current address: Department of Plant Sciences
and Plant Pathology, Montana State University, Bozeman, MT, USA; *These authors contributed equally to the work.
Microbes are estimated to comprise more than 90% of the biomass in the world’s oceans, are major drivers of biogeochemical cycles, and have turnover rates ranging from
hours to days. Despite the central role that microbes play in marine ecosystems, there is no robust method to evaluate taxon-specific mortality rates. Here, we report a method
that employs extracellular free-ribosomes as a proxy to evaluate taxon-specific microbial mortality. The method was validated with laboratory cultures of the marine
heterotrophic bacterium Vibrio natriegens strain PWH3a and the photoautotroph Synechococcus strain DC2, with and without grazers or viruses, to identify the origin and fate
of the extracellular free-ribosomes. Our results showed that both viral lysis and programmed-cell-death (PCD) contribute to free-ribosome production. Ribosomes were not
released when cells were grazed, but grazers could consume free-ribosomes. We show that extracellular free-ribosomes can be used to evaluate microbial mortality caused by
viral lysis and PCD.
This approach was applied to environmental samples by examining the taxonomic composition and relative abundance of free 16S-ribosomes in seawater samples collected
from the Strait of Georgia, British Columbia, Canada. Based on the presence of free ribosomes, lysis was detected in 1468 out of 3261 prokaryotic taxa, representing 17
bacterial and two archaeal phyla. Of these, lysis of 75 taxa could be detected in all ten samples. Based on the ratio of free ribosomes to cellular ribosomes, some taxa
associated with specific ecological niches appeared to be subject to high rates of lysis, including the genera Achromobacter, Alteromonas, Aureispira, Colwellia, Delftia,
Glaciecola, Methylobacterium, Owenweeksia, Pseudospirillum, Phyllobacterium, Roseobacter, SAR11, SAR92, Sufflavibacter and Synechococcus. Our results showed high-lysis
coupled with low-abundance, suggesting that taxa in lower abundance are subject to higher relative rates of cell lysis, consistent with previous suggestions. The ability to
estimate taxon-specific mortality as the result of cell lysis adds an important tool in our quest to explain the distribution and abundance of specific microbial taxa in nature.
1. Beginning of the idea
2. Ribsome production during viral lysis
 Sometimes, high-concentrations of
rRNA can be detected in metagenomic
samples for RNA viruses.
Is this RNA from ribosomes
released as the result of viral lysis?
If so, can these ribosomes be used to
provide taxon-specific estimates of
microbial mortality?
Cell lysis via
virus or PCD
1.00E+09
1.00E+08
1.00E+07
Copies/mL
Bacterial cell
35.6% of the sequences were from the phylum Proteobacteria
1.00E+06
1.00E+05
RT
1.00E+04
No RT
1.00E+03
Ribosomes
 High concentrations of
free ribosomes also occur
in uninfected cultures.
Why?
1.00E+02
1.00E+01
1.00E+00
Infected Culture
(10e8 cells)
Uninfected
Culture (10e8
cells)
Stationary Culture
(10e9+ cells)
Production of ribosomes by Vibrio
natriegens alone or infected with the phage
P1 as measured by qPCR.
Prokaryotes
3. Ribosome production during
bacterial growth
 Free ribosomes are produced by uninfected Vibrio
but not by uninfected Synechococcus.
4. Ribosome production during
grazing
 No ribosomes are released when cells are grazed
 Grazers consume extracellular free-ribosomes
Protist
Is this caused by PCD?
A
Bacteria - Vibrio
A
 Lysis was detected in 1468 out of 3261 prokaryotic taxa, from 10
environmental samples, representing 17 bacterial and 2 archaeal phyla
1.00E+10
The concentration of free
ribosomes is increased by
100-fold when cells are
infected by viruses
5. Taxon-specific lysis estimation
B
16S Ribosome - Vibrio
B
C
Bacteria - Synechococcus
C
16S Ribosome - Synechococcus
D
Bacterial (A) and protist (B) counts during the growth of V. natriegens
alone (open circles) or with the grazer P. bandaiensis (open circles).
The exracellular free-ribosomes (C) were measured at the same time.
Growth of bacterial strains (the heterotroph Vibrio natriegens strain
PWH3a, A and the photoautotroph Synechococcus strain DC2, C) and
production of ribosomes (B and D, respectively).
3.1. TEM of PCD
Apoptotic-like and Necrotic-like PCD appears to occur
in V. natriegens
6. Relationships between lysis, relative
abundance and growth rate
B
A
Lysis
Lysis
75 prokaryotic taxa with measureable lysis (extracellular free-ribosomes /
cellular-ribosomes) detected in all10 environmental samples from Quadra
staion#24 (0, 5, 30, 100, 265m) during and after spring-bloom in 2015.
1
D
C
Lysis
TAKE HOME MESSAGE
Free-ribosomes can be used to evaluate taxon-specific microbial
mortality caused by viral lysis and programmed-cell-death.
Lysis
Growth
-3
Vibrio natriegens strain PWH3a, TEM conducted by C. Deeg
Contact: [email protected];
[email protected]; [email protected]
 High-lysis coupled with low-abundance and low growth rate
(p < 2.2e-16)
High-lysis results in low abundance.
The taxa in low-abundance are mostly those with low
growth rates (A).
Acknowledgement
This work is supported by the Tula foundation and the Hakai Institute. We thank
Alvin Tian for assistance in sequences analysis and Chris Deeg for the TEM and
providing P. bandaiensis. As well, thanks to members of the Suttle Lab for many
suggestions, and for team members from the Hakai Research institute for
sample collection.