Comparative genomics and super-resolution microscopy of novel cyanophages reveals genomic isomerism, auxiliary genes, and a non-synchronous infection process
Abstract
Background: Microbial communities of multicellular, heterocyst-forming cyanobacteria assimilate organic nitrogen from dinitrogen gas through nitrogen fixation and are important photosynthetic primary producers. This makes them ecologically significant, particularly in freshwater, wetlands, and agricultural ecosystems. Cyanophages, viruses that specifically infect cyanobacteria play a key role in lysing them, terminating cyanobacterial blooms, and turning over the nutrients accumulated by these communities, with tremendous relevance for the biogeochemical cycles in these environments. However, only a limited number of cyanophages infecting N ₂ -fixing cyanobacteria have been genome sequenced to date, and it has remained unknown how their interaction proceeds with the individual cells within the filaments forming the backbone of these communities.
Results: We isolated and characterized three Caudoviricetes cyanophages (A-Lf14, A-Alj1, and A-Hlh1) specifically infecting the N ₂ -fixing cyanobacterium Anabaena sp. PCC 7120. Super-resolution microscopy revealed heterogeneous infection outcomes among adjacent cells within individual host filaments, indicating asynchronous infection progression at the single-cell level. Comparative genomics revealed a conserved ~30 kb invertible genomic region flanked by inverted repeats, a structural configuration with limited precedent in cyanophages. Using diagnostic PCR, Sanger sequencing, and read-mapping, we validated that two isomers stably coexist within phage populations. In contrast to the previously characterized cyanophage A-1(L), the three new phages encode a non-cyanobacterial alkaline phosphatase (PhoD). The expression of phoD was highly upregulated during infection, suggesting a phage-driven mechanism for phosphorus acquisition. Conversely, these phages lack the early-expressed tnpB endonuclease gene carried by A-1(L). Transcription of tnpB was strongly upregulated for both strands ( tnpB fwd and tnpB rev ) during A-1(L) infection, while infection by the three novel phages modestly induced the transcription of five identical host-encoded tnpB copies. In a highly unusual setting, both tnpB strands are protein-coding over their entire length, leading to the expression of a 347-residue protein from the tnpB rev strand.
Conclusions: This study reveals that genomic isomerism is a source of structural plasticity in freshwater cyanophages and uncovers functionally distinct auxiliary gene repertoires that link phage infection strategies to nutrient status. Our findings provide mechanistic insight into how cyanophages diversify their genomes, interact with their hosts and show that a phage gene can be protein-coding on both strands.
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BibTeX
@article{shi2026comparative,
title = {{Comparative genomics and super-resolution microscopy of novel cyanophages reveals genomic isomerism, auxiliary genes, and a non-synchronous infection process}},
author = {Yue Shi and Marcus Ziemann and Yang Zhao and Viktoria Reimann and Tao Zhu and Wolfgang R. Hess and Xuefeng Lu},
journal = {Microbiome},
year = {2026},
month = oct,
doi = {10.1186/s40168-026-02551-1},
url = {https://link.springer.com/10.1186/s40168-026-02551-1},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Comparative genomics and super-resolution microscopy of novel cyanophages reveals genomic isomerism, auxiliary genes, and a non-synchronous infection process
AU - Yue Shi
AU - Marcus Ziemann
AU - Yang Zhao
AU - Viktoria Reimann
AU - Tao Zhu
AU - Wolfgang R. Hess
AU - Xuefeng Lu
PY - 2026
DA - 2026/10/09
JO - Microbiome
DO - 10.1186/s40168-026-02551-1
UR - https://link.springer.com/10.1186/s40168-026-02551-1
AB - Background: Microbial communities of multicellular, heterocyst-forming cyanobacteria assimilate organic nitrogen from dinitrogen gas through nitrogen fixation and are important photosynthetic primary producers. This makes them ecologically significant, particularly in freshwater, wetlands, and agricultural ecosystems. Cyanophages, viruses that specifically infect cyanobacteria play a key role in lysing them, terminating cyanobacterial blooms, and turning over the nutrients accumulated by these communities, with tremendous relevance for the biogeochemical cycles in these environments. However, only a limited number of cyanophages infecting N ₂ -fixing cyanobacteria have been genome sequenced to date, and it has remained unknown how their interaction proceeds with the individual cells within the filaments forming the backbone of these communities. Results: We isolated and characterized three Caudoviricetes cyanophages (A-Lf14, A-Alj1, and A-Hlh1) specifically infecting the N ₂ -fixing cyanobacterium Anabaena sp. PCC 7120. Super-resolution microscopy revealed heterogeneous infection outcomes among adjacent cells within individual host filaments, indicating asynchronous infection progression at the single-cell level. Comparative genomics revealed a conserved ~30 kb invertible genomic region flanked by inverted repeats, a structural configuration with limited precedent in cyanophages. Using diagnostic PCR, Sanger sequencing, and read-mapping, we validated that two isomers stably coexist within phage populations. In contrast to the previously characterized cyanophage A-1(L), the three new phages encode a non-cyanobacterial alkaline phosphatase (PhoD). The expression of phoD was highly upregulated during infection, suggesting a phage-driven mechanism for phosphorus acquisition. Conversely, these phages lack the early-expressed tnpB endonuclease gene carried by A-1(L). Transcription of tnpB was strongly upregulated for both strands ( tnpB fwd and tnpB rev ) during A-1(L) infection, while infection by the three novel phages modestly induced the transcription of five identical host-encoded tnpB copies. In a highly unusual setting, both tnpB strands are protein-coding over their entire length, leading to the expression of a 347-residue protein from the tnpB rev strand. Conclusions: This study reveals that genomic isomerism is a source of structural plasticity in freshwater cyanophages and uncovers functionally distinct auxiliary gene repertoires that link phage infection strategies to nutrient status. Our findings provide mechanistic insight into how cyanophages diversify their genomes, interact with their hosts and show that a phage gene can be protein-coding on both strands.
SN - 2049-2618
ER -