Symbioses play a central role in shaping host development and physiology, yet how multiple symbiotic partners collectively influence complex biological processes remains poorly understood. In the fungal holobiont Rhizopus microsporus, bacterial (Mycetohabitans) and viral (narnaviruses) endosymbionts contribute to the formation of zygospores between compatible sexual partners, although their influence on early sexual reproduction remains largely unexplored. Here, we used mRNA sequencing data to evaluate the impact of bacterial and viral symbionts on the fungal transcriptional landscape at the onset of sexual development, revealing a substantial transcriptional reprogramming. Through network analysis, we identified candidate hub genes, including transcription factors, protein kinases, and signal transduction components, that may contribute to this developmental transition. By integrating sRNA-sequencing data, we found small-RNA signals correlating with early sexual success. Furthermore, we uncovered distinct processing patterns for the RmNV-20S and RmNV-23S viral symbionts, revealing that RmNV-23S processing shifted during the initiation of fungal sexual reproduction in the absence of the bacterial partner. Target prediction analyses further suggest that viral-derived sRNAs could influence distinct functional signatures in the fungal genome. Moreover, predicted interactions between endogenous or viral sRNAs and fungal transcripts suggest a potential role for sRNAs in fine-tuning the host transcriptional profile. Altogether, our findings provide a novel framework for understanding how multi-kingdom symbioses modulate host development and identify potential molecular factors that may contribute to the onset of sexual reproduction in R. microsporus.
Zobellella denitrificans MW1 is a halotolerant, facultatively anaerobic bacterium with high poly(3-hydroxybutyrate) (PHB) productivity that lacked a complete genome sequence. Here we report its first complete circular genome (4.10 Mb, 63.6% GC), obtained by hybrid Illumina–Nanopore assembly, with multi-scale cultivation data. The genome encodes a conserved phaCAB operon, multiple sucrose uptake and catabolic systems, and a canonical Pho regulon with polyphosphate metabolism genes (ppk, ppx). Ten CRISPR arrays, an intact prophage and no acquired multidrug resistance determinants were identified. Carbohydrate utilization was broad yet selective: sucrose, glucose and fructose supported the highest biomass, sucrose yielding up to 76% PHB of cell dry weight (CDW) in flasks; sucrose-rich beet molasses sustained PHB accumulation in flasks and 5 L bioreactors. In single, non-replicated fed-batch runs, a high-phosphate process reached 23 g L⁻¹ CDW with minimal PHB, whereas a low-phosphate process reached 53 g L⁻¹ CDW with 78% PHB. Because each regime was run once, these data are observations rather than a statistically supported comparison. They suggest that phosphate availability may be an important variable governing the growth-versus-storage balance, and that sucrose-rich industrial side-streams can be converted into PHB. The genomic co-occurrence of the Pho regulon, polyphosphate metabolism and phaCAB offers a hypothesis that remains to be tested by expression, enzyme-activity and targeted genetic experiments. Overall, MW1 is established as a genomically resolved platform for PHB production from low-cost, sucrose-rich feedstocks under saline conditions.
Mercury (Hg) is a widespread contaminant that poses significant risks to human health, mainly through dietary exposure to methylmercury (MeHg). Strategies to reduce Hg bioaccesibility in the gastrointestinal tract are therefore of increasing interest. In this study, a probiotic bacterium, Lacticaseibacillus paracasei, was engineered to express different combinations of mer genes involved in Hg detoxification. Recombinant strains were evaluated for resistance to inorganic Hg [Hg(II)] and MeHg, as well as for their ability to remove these species from culture media. Expression of mer genes enhanced Hg resistance of the L. paracasei cells, with the strain carrying the full merETPAB1 gene set showing the highest tolerance. In contrast, expression of merB1 without merA increased MeHg-induced growth inhibition, indicating the toxicity of intracellularly generated Hg(II). Strains co-expressing merA and merB1 efficiently removed MeHg from culture media, achieving up to 93% decrease, likely via its reduction to volatile elemental mercury [Hg(0)], whereas Hg(II) removal was limited. MeHg removal by an L. paracasei strain expressing merETPAB1 was also observed in the presence of human intestinal microbiota under anaerobic conditions. However, when applied to the bioaccessible fraction of simulated digests of Hg-contaminated fish (Makaira nigricans), Hg removal was lower (20–40%) compared with pure MeHg conditions.These results provide proof of concept that genetically engineered lactic acid bacteria expressing mer genes can reduce Hg contents in food within simulated gastrointestinal environments. This approach represents a promising strategy to mitigate dietary exposure to Hg, although further optimization and in vivo validation are required.
The aerobic electron transport chain (ETC) is a major determinant of metabolic flexibility and ecological success in Pseudomonas, yet the evolutionary dynamics shaping its regulatory architecture remain poorly understood. Here we combine phylogenomics, promoter analysis, and functional assays across 249 Pseudomonas species to reveal extensive diversification in ETC composition and in the regulatory control of a key aerobic terminal oxidase, the Cyo ubiquinol oxidase. Cyo was detected in 83.5% of strains, and the regulatory regions controlling its expression displayed remarkable divergence. Promoter sequences upstream of cyoA segregated into five major classes with low or no similarity between groups. Reporter assays demonstrated that this promoter diversification translates into distinct regulatory behaviours. When cultured in a rich medium, Pseudomonas putida KT2440 exhibited a sharp early-exponential induction of cyoA, whereas Pseudomonas fluorescens, Pseudomonas syringae, Pseudomonas rhizosphaerae, and Pseudomonas aeruginosa showed a much lower induction. In a defined growth medium, Cyo activation varied strongly depending on the carbon source. These species-specific patterns were independent of growth rate, highlighting that regulatory evolution, rather than metabolic demand alone, drives Cyo expression dynamics. Loss of Cyo in P. putida reduced growth rate up to 37% and triggered broad transcriptional reprogramming involving respiration, carbon assimilation, and stress responses. Notably, lack of Cyo led to a > 13-fold increase in the expression of the CIO ubiquinol oxidase. Together, these results show that both the composition and regulatory control have undergone extensive evolutionary diversification in Pseudomonas. This regulatory rewiring enables lineage-specific respiratory behaviours that likely contribute to ecological adaptation.
Ferric reductases are oxidoreductases that catalyze the reduction of environmental Fe³ ⁺ to bioavailable Fe² ⁺ and mediate cellular iron uptake and metabolism. However, the specific biological functions of ferric reductases in Colletotrichum gloeosporioides have not been fully characterized. In this study, 15 ferric reductase-like proteins were screened from the genome annotation of C. gloeosporioides, and CgFre2 was selected for further functional characterization. Targeted deletion of CgFre2 resulted in reduced mycelial growth and a sharp decrease in conidiation. Further observation of conidial development showed delayed conidial germination and an increased incidence of abnormal appressorium formation, which impaired the proper morphogenesis of infection structures and thereby significantly attenuated the pathogenicity of the strain on chili peppers. Moreover, the expression levels of cell wall-related genes were downregulated in the CgFre2 deletion mutant, which exhibited impaired cell wall integrity and hypersensitivity to oxidative stress. Meanwhile, the expression levels of ferric reductase-like genes CgFre3, CgFre6, CgFre11, CgFre13, and CgFre15 were markedly upregulated in the CgFre2 mutant, whereas ferric reductase activity was reduced to 48% of the WT level. Notably, exogenous Fe³ ⁺ supplementation restored infection structure formation and pathogenicity on pepper leaves. Collectively, these findings reveal the biological function of ferric reductase CgFre2 and provide a theoretical basis for developing iron‑targeted green control strategies against pepper anthracnose.
