Genome-resolved characterization of candidate thermotolerance traits and predicted protein conformational behavior in Calditerricola during hyperthermophilic composting of organic wastes.

Bacterial Community and Predicted Functional Gene Dynamics Relevant to the Potential Biodegradation of Polybutylene Adipate Terephthalate (PBAT)

Polybutylene adipate terephthalate (PBAT) is a widely used biodegradable plastic, but its microbial degradation mechanisms and environmental responses remain poorly understood. In this study, two compost inocula (K and I) were used to establish PBAT enrichment cultures under mesophilic (35 °C, M) and thermophilic (58 °C, T) conditions, and the effects of temperature and inoculum source on bacterial community structure and PICRUSt2-derived predicted functional profiles were investigated. Surface analyses revealed cracking, erosion, and structural collapse of PBAT under all conditions, with more pronounced degradation in compost I-derived cultures. Microbial diversity decreased during enrichment, accompanied by dominance of specific genera. Under mesophilic conditions, Pseudoxanthomonas dominated (72.6%–91.0%) and showed strong positive correlations with predicted ester bond hydrolysis-related KOs (p < 0.05), suggesting a potential association with initial hydrolysis. Under thermophilic conditions, compost K cultures were dominated by Thermoflavifilum and Rhodothermus, which were mainly associated with predicted hydrolysis-related KOs, whereas compost I cultures were dominated by Thermoflavifilum and Thermopolyspora and showed stronger associations with predicted KOs related to adipate and terephthalic acid metabolism. Predicted aromatic intermediate metabolism-related KOs, including K04101, K01055, and K01607, were also higher in compost I-derived thermophilic cultures. Overall, temperature and inoculum composition jointly shaped bacterial community assembly and predicted PBAT degradation-related functional potential. Thermophilic conditions combined with compost I were associated with functionally differentiated bacterial communities and higher predicted potential for downstream PBAT-derived intermediate metabolism. These findings provide a microbial ecological basis for temperature- and inoculum-guided enrichment strategies to improve PBAT biodegradation and biodegradable plastic waste treatment.

Experimental, Genomic, and Structural Evidence Supporting Putative PET‐Hydrolases in Thermophilic Bacteria Isolated From Hot Springs in Cajamarca, Peru

ABSTRACT Polyethylene terephthalate (PET) waste represents a major environmental challenge due to limited recycling solutions. Thermophilic bacteria from geothermal environments harbor diverse enzymatic machinery adapted to extreme conditions, offering promising biocatalysts for plastic degradation; however, biological resources from Peru and other South American countries remain scarce. We characterized four bacterial strains isolated from two geothermal sites in Cajamarca, Peru, screened for PET hydrolysis at 50°C. Whole‐genome sequencing using hybrid assembly achieved near‐complete circular genomes. GTDB‐Tk classification identified three species: Neobacillus thermocopriae (strain 19A), Bacillus licheniformis (strains 16P and BI2), and Brevibacillus agri (strain BI8). Quantitative assays revealed that strain 16P achieved the highest mass loss (0.598%), followed by strain BI8 (0.449%). ATR‐FTIR analysis of the incubated sheets showed a significant reduction of the ester carbonyl index in strains 16P, 19A, and BI8 relative to both non‐incubated PET and an abiotic control, whereas strain BI2 did not differ from the controls, indicating preferential modification of ester bonds at the sheet surface. Genome mining and structure‐based homology searches identified multiple candidate enzymes similar to validated PETases and carboxylesterases, including PETase46‐like homologs in strains BI8 and 16P and a terephthalate‐active carboxylesterase homolog in strain 16P. Molecular docking supported the conservation of catalytic geometry and substrate‐binding sites in these candidates. This work represents one of the first systematic genomic and structural characterizations of putative PET‐hydrolases in Peruvian geothermal bacteria, expanding knowledge of extremophile diversity and advancing thermostable enzymes for sustainable plastic waste management.

Physiological and genomic characterization of Methanothermobacter thermautotrophicus strain Z-73A, an isolate from deep underground.

Underground biomethanation represents a promising solution for green energy storage. An isolate of Methanothermobacter thermautotrophicus was obtained in previous study from an underground gas reservoir where it played a key role in a successful in-situ biomethanation experiment. In this study, the physiological and environmental tolerances of the strain are characterised. Optimal methane evolution rates were observed at 55-65 °C and pH 6.5-7.0, methane production was significantly increased by agitation (up to 2.29 mmol CH4·h-1·L-1). The strain showed higher tolerance to NH4Cl (up to 0.6 M) than to NaCl (optimal 0.04-0.4 M). The strain was also tested for its resistance to silver nanoparticles. The whole-genome sequence of the strain was determined, and the corrosion potential was evaluated. To visualise the cells, Scanning Electron Microscopy (SEM) and a method of Advanced Environmental Scanning Electron Microscopy (A-ESEM) were used. These findings describe the behaviour of M.thermautotrophicus under suboptimal conditions and highlight the need for further optimization to improve methane production yields.

From phenotype to mechanism: background-dependent function of CtDnaJ.16 in thermotolerance-divergent Coriolopsis trogii haploids

The two haploids, Ct001_29 and Ct001_31, derived from the thermotolerant strain Coriolopsis trogii Ct001, differ in mating type and exhibit distinct thermotolerance capacities. To investigate the underlying molecular mechanisms, their phenotypic responses to different temperatures were characterized, and comparative RNA-sequencing (RNA-Seq) analyses were performed using an upgraded, manually curated genome annotation. Phenotypic observations revealed that Ct001_31 displayed more robust hyphal growth at 35 °C than Ct001_29, including denser mycelia, larger hyphal diameter, smoother hyphal surfaces, and the ability to sustain growth at 40 °C. Through manual correction, a high-quality genome annotation for Ct001_31 was generated, comprising 12,953 high-confidence genes. Comparative RNA-Seq analysis identified 108 up-regulated and 121 down-regulated genes commonly shared between the two haploids at 35 °C. The up-regulated genes in Ct001_29 were significantly enriched in functions related to protein folding and heat response, whereas no significant functional enrichment was observed among the differentially expressed genes (DEGs) in Ct001_31 at 35 °C. Notably, CtDnaJ.16 showed increased expression in Ct001_31 at 35 °C, while no significant change was observed in Ct001_29. Functional investigation revealed that Ct31DnaJ.16 was specifically involved in thermotolerance in Ct001_31, with transformants overexpressing this gene capable of germinating at 43 °C. In contrast, neither overexpression nor RNA interference of Ct29DnaJ.16 in Ct001_29 led to detectable phenotypic alterations. These findings demonstrate that CtDnaJ.16 plays a strikingly haploid-specific role in thermotolerance, deepening our understanding of functional differentiation between allelic genes.

Proteomics analysis of Pseudomonas plecoglossicida under thermal stress: the Entner-Doudoroff pathway as a temperature-dependent metabolic switch for thermal adaptation

Genomic insights from a long-read assembly of a deep-sea Chromohalobacter israelensis strain from Santos Basin pockmarks field

Analysis revealed genes for metabolizing diverse carbohydrate sources, a complete ectoine synthesis cluster essential for halophily, and genes conferring tolerance to osmotic stress, low temperatures, and pH, underscoring its polyextremophilic capacity.