Analysis of metagenomic data
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Temperature-driven nitrogen mixotrophy shapes marine cyanobacteria Prochlorococcus and Synechococcus latitudinal distribution pattern
Temperature, photosynthetically active radiation, and nutrient availability are pivotal in determining the global distribution of marine cyanobacteria. This study demonstrates that the distinct latitudinal niches of Synechococcus and Prochlorococcus are modulated by their temperature-dependent, mixotrophic nitrogen utilization strategies. Comparative genomic analysis reveals that Prochlorococcus ecotypes differ in their nitrogen transporter repertoire, with low-light ecotypes harboring ~12 transporters, while high-light ecotypes possess ~5 transporters. Conversely, Synechococcus displays greater genomic flexibility, with 17 nitrogen transporters. Reanalysis of Tara Oceans metatranscriptomic data identifies ~15 °C as a key thermal threshold for cyanobacterial distribution. In warmer regions (15-30°C, 35˚S-40˚N), Prochlorococcus optimizes nitrogen transporter functions across their ecotypes, whereas Synechococcus, leveraging its broad nitrogen transporters, exhibits functional plasticity, utilizing inorganic nitrogen in warmer waters and organic nitrogen in cooler waters ranging from -2°C to 15 °C beyond 35˚S/40˚N. Our findings underscore the critical role of temperature-driven mixotrophic nitrogen utilization in shaping the biogeographical patterns of marine cyanobacteria.
Analysis of microbial composition and sharing in low-biomass human milk samples: a comparison of DNA isolation and sequencing techniques
Human milk microbiome studies are currently hindered by low milk bacterial/human cell ratios and often rely on 16S rRNA gene sequencing, which limits downstream analyses. Here, we aimed to find a method to study milk bacteria and assess bacterial sharing between maternal and infant microbiota. We tested four DNA isolation methods, two bacterial enrichment methods and three sequencing methods on mock communities, milk samples and negative controls. Of the four DNA isolation kits, the DNeasy PowerSoil Pro (PS) and MagMAX Total Nucleic Acid Isolation (MX) kits provided consistent 16S rRNA gene sequencing results with low contamination. Neither enrichment method substantially decreased the human metagenomic sequencing read-depth. Long-read 16S-ITS-23S rRNA gene sequencing biased the mock community composition but provided consistent results for milk samples, with little contamination. In contrast to 16S rRNA gene sequencing, 16S-ITS-23S rRNA gene sequencing of milk, infant oral, infant faecal and maternal faecal DNA from 14 mother-infant pairs provided sufficient resolution to detect significantly more frequent sharing of bacteria between related pairs compared to unrelated pairs. In conclusion, PS or MX kit-DNA isolation followed by 16S rRNA gene sequencing reliably characterises human milk microbiota, and 16S-ITS-23S rRNA gene sequencing enables studies of bacterial transmission in low-biomass samples.
Gut microbiota-derived hexa-acylated lipopolysaccharides enhance cancer immunotherapy responses
The gut microbiome modulates immunotherapy treatment responses, and this may explain why immune checkpoint inhibitors, such as anti-PD-1, are only effective in some patients. Previous studies correlated lipopolysaccharide (LPS)-producing gut microbes with poorer prognosis; however, LPS from diverse bacterial species can range from immunostimulatory to inhibitory. Here, by functionally analysing faecal metagenomes from 112 patients with melanoma, we found that a subset of LPS-producing bacteria encoding immunostimulatory hexa-acylated LPS was enriched in microbiomes of clinical responders. In an implanted tumour mouse model of anti-PD-1 treatment, microbiota-derived hexa-acylated LPS was required for effective anti-tumour immune responses, and LPS-binding antibiotics and a small-molecule TLR4 antagonist abolished anti-PD-1 efficacy. Conversely, oral administration of hexa-acylated LPS to mice significantly augmented anti-PD-1-mediated anti-tumour immunity. Penta-acylated LPS did not improve anti-PD-1 efficacy in vivo and inhibited hexa-acylated LPS-induced immune activation in vitro. Microbiome hexa-acylated LPS therefore represents an accessible predictor and potential enhancer of immunotherapy responses.
Phylogenetically and metabolically diverse autotrophs in the world’s deepest blue hole
The world’s deepest yongle blue hole (YBH) is characterized by sharp dissolved oxygen (DO) gradients, and considerably low-organic-carbon and high-inorganic-carbon concentrations that may support active autotrophic communities. To understand metabolic strategies of autotrophic communities for obtaining carbon and energy spanning redox gradients, we presented finer characterizations of microbial community, metagenome and metagenome-assembled genomes (MAGs) in the YBH possessing oxic, hypoxic, essentially anoxic and completely anoxic zones vertically. Firstly, the YBH microbial composition and function shifted across the four zones, linking to different biogeochemical processes. The recovery of high-quality MAGs belonging to various uncultivated lineages reflected high novelty of the YBH microbiome. Secondly, carbon fixation processes and associated energy metabolisms varied with the vertical zones. The Calvin–Benson–Bassham (CBB) cycle was ubiquitous but differed in affiliated taxa at different zones. Various carbon fixation pathways were found in the hypoxic and essentially anoxic zones, including the 3-hyroxypropionate/4-hydroxybutyrate (3HP/4HB) cycle affiliated to Nitrososphaeria, and Wood-Ljungdahl (WL) pathway affiliated to Planctomycetes, with sulfur oxidation and dissimilatory nitrate reduction as primary energy-conserving pathways. The completely anoxic zone harbored diverse taxa (Dehalococcoidales, Desulfobacterales and Desulfatiglandales) utilizing the WL pathway coupled with versatile energy-conserving pathways via sulfate reduction, fermentation, CO oxidation and hydrogen metabolism. Finally, most of the WL-pathway containing taxa displayed a mixotrophic lifestyle corresponding to flexible carbon acquisition strategies. Our result showed a vertical transition of microbial lifestyle from photo-autotrophy, chemoautotrophy to mixotrophy in the YBH, enabling a better understanding of carbon fixation processes and associated biogeochemical impacts with different oxygen availability.
Phylogenetic and ecophysiological novelty of subsurface mercury methylators in mangrove sediments
Mangrove sediment is a crucial component in the global mercury (Hg) cycling and acts as a hotspot for methylmercury (MeHg) production. Early evidence has documented the ubiquity of well-studied Hg methylators in mangrove superficial sediments; however, their diversity and metabolic adaptation in the more anoxic and highly reduced subsurface sediments are lacking. Through MeHg biogeochemical assay and metagenomic sequencing, we found that mangrove subsurface sediments (20–100 cm) showed a less hgcA gene abundance but higher diversity of Hg methylators than superficial sediments (0–20 cm). Regional-scale investigation of mangrove subsurface sediments spanning over 1500 km demonstrated a prevalence and family-level novelty of Hg-methylating microbial lineages (i.e., those affiliated to Anaerolineae, Phycisphaerae, and Desulfobacterales). We proposed the candidate phylum Zixibacteria lineage with sulfate-reducing capacity as a currently understudied Hg methylator across anoxic environments. Unlike other Hg methylators, the Zixibacteria lineage does not use the Wood–Ljungdahl pathway but has unique capabilities of performing methionine synthesis to donate methyl groups. The absence of cobalamin biosynthesis pathway suggests that this Hg-methylating lineage may depend on its syntrophic partners (i.e., Syntrophobacterales members) for energy in subsurface sediments. Our results expand the diversity of subsurface Hg methylators and uncover their unique ecophysiological adaptations in mangrove sediments.
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