Proyash Roy

Aims: Bacteria experience various selective pressures from the environment, including exposure to antibiotics and bacteriophages, which shape their defence strategies and horizontal gene transfer (HGT) dynamics. The relationship between defence system repertoires and HGT in clinically relevant Acinetobacter species remains poorly understood, limiting our ability to predict resistance evolution and design targeted phage therapies. Methods and results: We analysed 132 genomes from 18 Acinetobacter species, focusing on the interplay between defence architectures and HGT markers. Our results reveal that defence repertoires differed across lineages. Most Acinetobacter spp. harbour multiple defence systems, whereas the clinically dominant A. baumannii international clone 2 carried fewer but was strongly enriched for the phosphorothioation-based SspBCDE system and had very few restriction–modification systems. Strikingly, many defence systems were rarely found together. Defence genes were frequently associated with the presence of mobile elements, antibiotics, and heavy metal resistance. Plasmid-borne defence systems, especially BREX, were prevalent, highlighting the role of mobile elements in distributing both anti-phage defence and clinically relevant resistance traits. Conclusions: Our results indicate that clinical success in A. baumannii is associated with a niche-driven defence profile and extensive linkage between defence genes, mobile elements, and resistance loci, which are likely to influence both HGT-mediated resistance acquisition and phage susceptibility.

2. Introduction: Bacteria face relentless selective pressure from both antibiotics and bacteriophages, driving the evolution of diverse defence systems and shaping horizontal gene transfer (HGT) dynamics. Gap Statement: The relationship between bacterial defence system repertoires and horizontal gene transfer in clinically relevant Acinetobacter species remains poorly understood, limiting our ability to predict resistance evolution and design targeted phage therapies. Aim: To characterize defence arsenal across Acinetobacter species and investigate their associations with different HGT markers Methodology: We performed a comparative genomic analysis of 132 genomes from 18 Acinetobacter species, focusing on the interplay between defence architectures and HGT markers. Results: Our result reveals pronounced differences in defence system repertoires between lineages: while most Acinetobacter spp. harbour a multilayered arsenal, the clinically dominant A. baumannii international clone 2 (IC2) exhibits a depleted but specialised defence landscape, notably enriched in the phosphorothioation-based SspBCDE system and nearly devoid of restriction-modification systems. Strikingly, many defence systems display mutual exclusivity, and defence system is tightly linked to the presence of mobile elements, antibiotics, and heavy metal resistance. Plasmid-borne defence systems, especially BREX, are prevalent, highlighting the role of mobile elements in distributing both immunity and resistance traits. These patterns suggest that successful clinical lineages finely balance immune defences and genetic plasticity, facilitating rapid adaptation under antimicrobial selection and phage threats. Conclusion: These findings provide insights into evolutionary trade-offs underpinning multidrug resistance and have implications for designing targeted phage therapies against recalcitrant Acinetobacter infections. 3. Data summary: All bioinformatics analysis workflows, custom scripts, and R code are publicly available at https://github.com/vikos77/Acinetobacter-defence-systems . The repository includes bash scripts for genome downloading and tool execution (scripts 1-10), R scripts for statistical analysis and visualization (scripts 1-9), and conda environment files for reproducible software installation. Complete analysis protocols are documented in the repository README and individual script headers. All software versions and parameters are specified in the code documentation. 4. Impact Statement: This study reveals how defence system composition in Acinetobacter species is shaped by ecological niche and linked to horizontal gene transfer, antibiotic resistance, and mobile genetic elements. Notably, the clinically dominant A. baumannii IC2 lineage exhibits a streamlined defence repertoire favouring genetic plasticity, facilitating rapid adaptation under antibiotic and phage pressure. These findings enhance our understanding of resistance evolution and support the rational design of phage-based therapies targeting multidrug-resistant pathogens.

Background: Despite the knowledge that the soil–plant–microbiome nexus is shaped by interactions amongst its members, very little is known about how individual symbioses regulate this shaping. Even less is known about how the agriculturally important symbiosis of nitrogen-fixing rhizobia with legumes is impacted according to soil type, yet this knowledge is crucial if we are to harness or improve it. We asked how the plant, soil and microbiome are modulated by symbiosis between the model legume Medicago truncatula and different strains of Sinorhizobium meliloti or Sinorhizobium medicae whose nitrogen-fixing efficiency varies, in three distinct soil types that differ in nutrient fertility, to examine the role of the soil environment upon the plant–microbe interaction during nodulation. Results: The outcome of symbiosis results in installment of a potentially beneficial microbiome that leads to increased nutrient uptake that is not simply proportional to soil nutrient abundance. A number of soil edaphic factors including Zn and Mo, and not just the classical N/P/K nutrients, group with microbial community changes, and alterations in the microbiome can be seen across different soil fertility types. Root endosphere emerged as the plant microhabitat more affected by this rhizobial efficiency-driven community reshaping, manifested by the accumulation of members of the phylum Actinobacteria. The plant in turn plays an active role in regulating its root community, including sanctioning low nitrogen efficiency rhizobial strains, leading to nodule senescence in particular plant–soil–rhizobia strain combinations. Conclusions: The microbiome–soil–rhizobial dynamic strongly influences plant nutrient uptake and growth, with the endosphere and rhizosphere shaped differentially according to plant–rhizobial interactions with strains that vary in nitrogen-fixing efficiency levels. These results open up the possibility to select inoculation partners best suited for plant, soil type and microbial community.

Legumes house nitrogen-fixing endosymbiotic rhizobia in specialized polyploid cells within root nodules, which undergo tightly regulated metabolic activity. By carrying out expression analysis of transcripts over time in Medicago truncatula nodules, we found that the circadian clock enables coordinated control of metabolic and regulatory processes linked to nitrogen fixation. This involves the circadian clock-associated transcription factor LATE ELONGATED HYPOCOTYL (LHY), with lhy mutants being affected in nodulation. Rhythmic transcripts in root nodules include a subset of nodule-specific cysteine-rich peptides (NCRs) that have the LHY-bound conserved evening element in their promoters. Until now, studies have suggested that NCRs act to regulate bacteroid differentiation and keep the rhizobial population in check. However, these conclusions came from the study of a few members of this very large gene family that has complex diversified spatio-temporal expression. We suggest that rhythmic expression of NCRs may be important for temporal coordination of bacterial activity with the rhythms of the plant host, in order to ensure optimal symbiosis.

Here, we report an improved and complete genome sequence of Sinorhizobium (Ensifer) meliloti strain WSM1022, a microsymbiont of Medicago species, revealing its tripartite structure. This improved genome sequence was generated combining Illumina and Oxford nanopore sequencing technologies to better understand the symbiotic properties of the bacterium. The 6.75 Mb WSM1022 genome consists of three scaffolds, corresponding to a chromosome (3.70 Mb) and the pSymA (1.38 Mb) and pSymB (1.66 Mb) megaplasmids. The assembly has an average GC content of 62.2% and a mean coverage of 77X. Genome annotation of WSM1022 predicted 6058 protein coding sequences (CDSs), 202 pseudogenes, 9 rRNAs (3 each of 5S, 16S, and 23S), 55 tRNAs, and 4 ncRNAs. We compared the genome of WSM1022 to two other rhizobial strains, closely related Sinorhizobium (Ensifer) meliloti Sm1021 and Sinorhizobium (Ensifer) medicae WSM419. Both WSM1022 and WSM419 species are high-efficiency rhizobial strains when in symbiosis with Medicago truncatula, whereas Sm1021 is ineffective. Our findings report significant genomic differences across the three strains with some similarities between the meliloti strains and some others between the high efficiency strains WSM1022 and WSM419. The addition of this high-quality rhizobial genome sequence in conjunction with comparative analyses will help to unravel the features that make a rhizobial symbiont highly efficient for nitrogen fixation.

Legumes house nitrogen-fixing endosymbiotic rhizobia in specialized polyploid cells within root nodules, which are factories of metabolic activity. We discovered that the circadian clock-associated transcriptional factor LATE ELONGATED HYPOCOTYL (LHY) affects nodulation in Medicago truncatula . By carrying out expression analysis of transcripts over time in nodules we found that the clock enables coordinated control of metabolic and regulatory processes linked to nitrogen fixation. Rhythmic transcripts in root nodules include a subset of Nodule-specific Cysteine Rich peptides (NCRs) that have the LHY-bound conserved Evening Element in their promoters. Until now, studies have suggested that NCRs act to regulate bacteroid differentiation and keep the rhizobial population in check. However, these conclusions came from the study of a few members of this very large gene family that has complex diversified spatio-temporal expression. We suggest that rhythmic expression of NCRs may be important for temporal coordination of bacterial activity with the rhythms of the plant host, in order to ensure optimal symbiosis. Highlights: The circadian clock-associated transcriptional factor LATE ELONGATED HYPOCOTYL (LHY) impacts on successful Medicago truncatula -rhizobial symbiosis The plant clock coordinates rhythmic patterns of metabolic and regulatory activity in nodules and drives rhythmic expression of a subset of Nodule-specific Cysteine Rich (NCR) genes. Rhythmic expression of NCRs may be important for temporal coordination of bacterial activity with plant host rhythms to ensure optimal symbiosis.

Legume-rhizobium symbiosis represents one of the most successfully co-evolved mutualisms. Within nodules, the bacterial cells undergo distinct metabolic and morphological changes and differentiate into nitrogen-fixing bacteroids. Legumes in the inverted repeat lacking clade (IRLC) employ an array of defensin-like small secreted peptides (SSPs), known as nodule-specific cysteine-rich (NCR) peptides, to regulate bacteroid differentiation and activity. While most NCRs exhibit bactericidal effects in vitro, studies confirm that inside nodules they target the bacterial cell cycle and other cellular pathways to control and extend rhizobial differentiation into an irreversible (or terminal) state where the host gains control over bacteroids. While NCRs are well established as positive regulators of effective symbiosis, more recent findings also suggest that NCRs affect partner compatibility. The extent of bacterial differentiation has been linked to species-specific size and complexity of the NCR gene family that varies even among closely related species, suggesting a more recent origin of NCRs followed by rapid expansion in certain species. NCRs have diversified functionally, as well as in their expression patterns and responsiveness, likely driving further functional specialisation. In this review, we evaluate the functions of NCR peptides and their role as a driving force underlying the outcome of rhizobial symbiosis, where the plant is able to determine the outcome of rhizobial interaction in a temporal and spatial manner.

Not available Dhaka Univ. J. Biol. Sci. 25(2): 201-205, 2016 (July)