Archives of Microbiology

ASS1 was isolated as a motile Stenotrophomonas strain from crude oil-contaminated soils in Tabasco, Mexico. We characterized this strain using physiological and biochemical traits. ASS1 grew at temperature 25-37 (optimally at 37 {degrees} C) and at pH 6 to 8 (optimally at pH 7 to 8). The assembled genome has a total length of 4.56MB with a G + C content of 66.6%. The 16S rRNA gene sequence analysis confirmed that this strain belongs to the genus Stenotrophomonas. Based on the 16S rRNA analysis, Stenotrophomonas geniculata ATCC 19374 is the closest species, and it shares 99.86% similarity with ASS1. Similarly, a phylogenomic tree based on core genome sequence revealed that the closest species to ASS1 is Stenotrophomonas geniculata ATCC 19374. The major fatty acids in ASS1 are C16:0, antesio C15:0, iso C12:0, iso C15:0, iso C17:0 and C18:0. The genome of ASS1 consists of 4,373,402 bp. The Average Nucleotide Identity (ANI) values for ASS1 which it shared with its closest phylogenetic neighbors, are Stenotrophomonas geniculata ATCC 19374 = JCM 13324 [T] 92.66 %, Stenotrophomonas maltophilia 13637[T] 92.15%, Stenotrophomonas maltophilia K279a 92.13% Stenotrophomonas maltophilia R551-3 92.15% Stenotrophomonas maltophilia MTCC 434 [T] 92.08% and Pseudomonas hibisicicola ATCC [T] 91.66%. ASS1 possesses genes that are essential for the degradation of Polycyclic Aromatic hydrocarbon. Genes such as 1, 2 dihydroxyl 1, 2 dihydronaphthalene dehydrogenase; MG068 17425, homologous to 2 hydroxyl chromene 2 carboxylate isomerases; MG 18055, homologous to salicylaldehyde dehydrogenase and MG068 20095, homologous to naphthalene 1, 2 dioxygenases were identified in ASS1. The dDDH value between ASS1 and its closest neighbor Stenotrophomonas geniculata ATCC 19374 = JCM 13324 [T] is 50%, which is the highest for all the typed species and as such we proposed that ASS1 is a novel species with the name Stenotrophomonas oleivorans sp. nov. sp. nov. and ASS1T as the typed strain

Carbapenem-resistant (CR) bacteria have emerged and been spreading beyond healthcare-associated facilities into the environment. It is recognized that toilet bowl water in patient rooms of healthcare-associated facilities can be one of internal reservoirs of CR bacteria. In accordance with this idea, toilet bowl water samples were collected from patient rooms in a tertiary healthcare-associated facility in North Macedonia, and meropenem (MEM)-resistant bacterial isolates were obtained from the toilet bowl water. In this study, because a MEM-resistant C. braakii isolate, that was one of MEM-resistant opportunistic pathogens, was obtained from the toilet water, whole-genome sequencing (WGS) of this isolate was performed to obtain genetic characteristics of the blaNDM-1-positive C. braakii isolate. By the WGS, four contigs were constructed, the longest contig, contig 1 (5,189,681 bp), contained blaCTX-M with some additional antimicrobial-resistance genes (ARGs). Interestingly, blaNDM-1 was detected in contig 2 (177,260 bp) and contig 3 (64,168 bp). Plasmid replicon of contig 2 was IncA/C2 but plasmid replicon of contig 3 was IncN and different from one of contig 2. Genetic structures surrounding blaNDM-1 were different between these two blaNDM-1-positive plasmids implying transfer or insertion of blaNDM-1 had occurred by IS or other mechanism. Further molecular epidemiology will be needed to explain the mechanism that allowed the C. braakii isolate to possess two structurally different blaNDM-1 plasmids.

Despite the successful cultivation of many microbes from rich bacterial communities inhabiting alkaline soda lakes, members of the bacterial phylum Verrucomicrobiota have so far been detected only through metagenomics. Here, we used alginate as a selective substrate to enrich and isolate two strains of haloalkaliphilic Verrucomicrobiota. The isolates share identical 16S rRNA gene sequences representing a new genus lineage, and, together with other metagenome assembled genomes, a new family within Opitutales. Cells of strains AB-alg1T (from soda lakes) and AB-alg4 (from soda solonchak soils) are small and motile cocci forming submerged colonies in soft alginate agar. They are saccharolytic heterotrophs growing aerobically on polysaccharides (alginate, starch and inulin) and sugars (glucose, fructose, mannose, sucrose, melezitose, maltose and cellobiose). They also grow anaerobically by fermentation of alginate and D-mannose and by coupling incomplete denitrification to oxidation of alginate. Both isolates are obligately alkaliphilic and moderately salt-tolerant. The dominant membrane phospholipids include phosphatidylcholines and diphosphatidylglycerols (cardiolipins). The genome of AB-alg1T features polysaccharide lyases of the PL6, 7, 15, 17, 38, and 39 families for depolymerization of alginate. Based on distinct phenotype and phylogeny, we propose classification of strains AB-alg1T (JCM 35393T=UQM 41574T) and AB-alg4 as Verruconatronum alginivorum gen. nov., sp. nov. within a new family Verruconatronumaceae. ImportanceThe presented isolates are the first isolated representatives of an environmental family of Opitutales, part of the core microbiome of alkaline soda lakes. These bacteria feed on polysaccharides. We present the key enzymatic machinery for the polysaccharide breakdown. These enzymes are high-pH tolerant and have potential for industry applications, for example in washing powders and biomass waste recycling.

Industrial waste containing hydrophobic pollutants, like oils and hydrocarbons, is toxic and difficult to degrade, posing both ecological and human health risks. Biosurfactants are eco-friendly surface-active compounds produced by microorganisms, known for their ability to lower surface and interfacial tension, enhancing the solubility and bioavailability of hydrophobic compounds, facilitating their breakdown. The current study focuses on isolating biosurfactant-producing bacteria from industrial waste sources near Dhaka, Bangladesh, and characterizing their properties, determining potential usage. Using diesel-enriched nutrient agar, bacterial strains were isolated and screened for biosurfactant production by oil displacement, emulsification index (E24%), and drop collapse assay. The most promising isolates were characterized according to their biochemical activities and 16S rRNA amplicon-based sequencing. Isolation and characterization of the surfactants have been carried out using chromatographic techniques. The identified bacteria passed the drop collapse and oil displacement tests. CTAB agar assay, indicates their anionic nature, showing an emulsification index ranging 10-41%. The potential biosurfactant producers belong to Bacillus, Pseudomonas, Acinetobacter, and Enterobacterium genera. The surfactants showed antibacterial, antifungal, and plant growth promotion activity and have been characterized in terms of pH stability, salinity, adhesion, and temperature tolerance. The study successfully identified and characterized potential biosurfactant-producing bacteria from industrial waste, highlighting their efficiency in breaking down hydrophobic pollutants and hydrocarbons. These microorganisms provide a green and economical substitute for synthetic surfactants due to their biodegradability and lower toxicity. Upon further research and scaling, these bacteria can be a good source of biosurfactants for potential applications in industrial, agricultural, and biomedical fields. IMPORTANCEThe study carries high significance as it creates multi-disciplinary scopes for utilizing these environmentally adapted biosurfactant-producing bacteria in industry, agriculture, and medicine. Since the bacterial isolates have hydrocarbon degradation ability, upon optimization for higher production, industrial usage in oil refinery and other industries can be adopted. Due to their biodegradable nature, usage in wound healing bandages and as antimicrobial agents in medicine will be noteworthy. The isolates have plant growth promotion ability and utilizing them as biofertilizer will reduce the dependency on chemical fertilizers. This is the first detailed report on biosurfactant-producing bacteria from this industrial waste-polluted Turag River of Dhaka City. Moreover, it compiles detailed screening protocols and methods for analyzing such environmentally friendly microbes. Such characterization also opens the scope for optimizing the production of the surfactant compounds on a large scale and utilizing them commercially.

This paper evaluated and compared the relative microalgal biomass accumulation of rocking, floating, and stationary bag photobioreactors. Microalga Neochloris oleoabundans was grown in these photobioreactors in batch mode for 24 days under illumination. The 50 L plastic bags (cell suspension volume 25 L) were placed on the surface of a rocking platform, an artificial pond or a stationary platform. In the pond, waves were generated by electrical fans which shake and mix microalgal cells within the plastic bags. The bags were supplied with 5% CO2 in air under elevated pressure inside of the bags. The rocking bag method significantly increased biomass yields to approximately 3-4 g * L-1, as compared to 0.16 g * L-1 in the floating photobioreactor and only 0.03 g * L-1 in the stationary type photobioreactor.

The Bacillus genus comprises physiologically versatile, endospore-forming bacteria widely distributed in natural environments. In this study, we report the isolation and genomic characterization of strain Bva_UNVM-123, recovered from agricultural soil in Pergamino, Argentina. Whole-genome sequencing using Illumina technology yielded a 5.1 Mbp draft genome assembled in 67 contigs with a GC content of 36%. Comparative genomic analyses using the TYGS server and digital DNADNA hybridization (dDDH) values supported its classification as a potentially novel species within the Bacillus sensu lato (s.l.) group. Genome annotation revealed 4,866 protein-coding genes, including multiple determinants conferring resistance to antibiotics (e.g., fosfomycin, tetracycline, beta-lactams) and toxic heavy metals (e.g., arsenic, cadmium, mercury), supporting its potential application in bioremediation. Additionally, PathogenFinder predicted a low probability of human pathogenicity (0.207), reinforcing its safety for environmental use. Functional classification based on Swiss-Prot further supported a metabolically versatile profile and revealed the presence of resistance-related categories associated with environmental adaptation. This study adds to the growing knowledge of environmental Bacillus species and their biotechnological potential

Currently, there are three recognized rhizobial genera belonging to the beta branch of the proteobacteria; Trinickia, Paraburkholderia, and Cupriavidus. These beta-rhizobia have been found associated with legume species mainly within the Mimosoideae and Papillonoideae. Most diversity, evolutionary, and functional studies have focused on Paraburkholderia, whereas few have addressed the diversity and evolution of symbiosis in the Cupriavidus genus. The present work aimed to provide an actual view of the symbiotic Cupriavidus diversity and to analyse the origin and evolution of their symbiotic genes. Using whole-genome information for phylogenetic reconstruction, we showed that the described symbiotic Cupriavidus strains belong to five distinct lineages, although they are intermixed with non-symbiotic species. The high synteny and sequence conservation of symbiotic genes suggest a common origin of acquisition for all rhizobial Cupriavidus described so far. However, we observed very low sequence conservation among (mega)plasmids carrying the symbiotic island, excluding the existence of a conserved symbiotic plasmid within beta-rhizobia. We can conclude that up to now there are five rhizobial species within the Cupriavidus genus, and we predict the description of new symbiotic species in the near future.

BackgroundGlobally, seagrass ecosystems are threatened by anthropogenic activities that are leading to increased levels of eutrophication, coastal pollution and thermal conditions. Consequently, there is a growing need to develop new approaches that work to mitigate these stressors and enhance restoration efforts in seagrass meadows. One promising strategy is to identify, isolate and characterize microbial consortia that are likely to support seagrass productivity. However, our current understanding of key microbial functions that support plant growth in marine systems is limited. Based on evidence from terrestrial plant-microbe systems, seagrass-associated bacteria are expected to provide the plant with nitrogen and phosphorus resources while detoxifying sulfur and producing phytohormones. Here, we sequenced 61 bacterial cultures isolated from the rhizosphere, rhizoplane, and endosphere of the seagrass, Zostera marina to identify a consortium of six putative plant growth promoting (PGP) candidates. ResultsOur cultivation approach using plant-based media allowed us to isolate 201 bacteria from Z. marina, which reflected 18% of the total microbial diversity of the starting inoculum. Genomic and phenotypic analyses of the 61-sequenced pure-cultures revealed that most of the sequenced taxa were able to mobilize nitrogen primarily through catabolic pathways, including denitrification (51%), dissimilatory nitrate reduction to ammonia (71%), and C-N bond cleavage (83%). Six of the isolates, which represent new lineages of Agarivorans, coded for the nitrogenase gene cassette. Additionally, 52% of the genomes had genes for sulfur and/or thiosulfate oxidation, 88.5% for phosphorus solubilization, and 60.5% for IAA production. Genomic analysis also revealed that some pathways, including denitrification and dissimilatory nitrite to ammonia DNRA, required cross-species cooperation as no one taxa contained all the genes needed to complete these metabolic pathways. Based on draft genome models and results from phenotypic assays, isolates Streptomyces sp. (Iso23 and Iso384), Mesobacillus sp (Iso127), Roseibuim sp. (Iso195), Peribacillus sp. (Iso49), and Agarivorans sp. (Iso311) represent a minimal microbial community that is likely to promote seagrass growth and enhance restoration efforts. ConclusionOur work provides a detailed genomic and phenotypic analysis of bacteria isolated from Z. marina and identifies a minimal microbial community with complementary PGP traits. Isolating, identifying and characterizing bacteria that promote seagrass growth is critical towards enhancing restoration efforts of seagrass meadows.

Nowadays, finding new sustainable ways to combat plastic pollution is a pressing challenge. Here, we provide a comprehensive genome mining analysis of 284 publicly available Stutzerimonas genomes for potential PET-active enzymes (PETases). While Stutzerimonas is a relatively newly established genus, it emerges as an interesting candidate in the search for novel biocatalysts. Hence, the first pangenome assessment of this genus based on its high-quality publicly available genomes was performed. An increasingly open pangenome was revealed, suggesting the versatility and adaptability of these strains to a variety of ecological niches. Moreover, functional characterisation of a new isolate, Stutzerimonas frequens VG-9, was carried out, confirming that enzymes found via in silico analyses may indeed display activity towards different polyesters. In summary, this study provides insights into the diversity of PETase homologues within still underexplored bacterial hosts, offering new perspectives for enzyme discovery in the Pseudomonadaceae family. Impact StatementMicrobial enzymes known as PETases have emerged as promising candidates for the biological degradation of PET. This study investigated the potential of underexplored bacterial genera by genome mining of PETase homologues. Our findings provide new insights into the distribution of PETase-like enzymes in the Pseudomonadaceae family, offering a more comprehensive view of their plastic degradation capacity. These results hold practical implications for the development of optimized enzyme discovery strategies, while also highlighting the vast genetic plasticity of Pseudomonadaceae. We also provided the first report on the Stutzerimonas pangenome and insights into the enzymatic activity towards polyesters of a newly isolated strain. Hence, the role of this genus as a highly adaptable and versatile entity was reinforced, further disclosing it as a potential source of novel biocatalysts. Data SummaryThe genome of S. frequens VG9 has been deposited in Genbank under the accession number SAMN49487720. The accession numbers of all analyzed genomes are listed in Tables S2 and S3 (available in the online Supplementary Material).

Non-pathogenic Xanthomonas (NPX) from a diverse plant hosts are being reported on an increasing basis. There are also reports of multiple species forming communities on a single host plant, such as rice, and, given their role as core endophytes in protecting plants from pathogens, it is essential to isolate and characterization of more NPX species from diverse host plants. Using phylogenomic analysis of publicly available Xanthomonas genome sequences, we identified a novel clade comprising NPX strains from diverse hosts. One of the strains previously reported from our lab is from healthy rice seeds and was reported to be non-pathogenic, with bio-protection function against the bacterial leaf blight pathogen. Genomic investigation confirmed the lack of type III secretion system and its effectors, consistent with their non-pathogenic nature. These strains also harbour core and unique biosynthetic loci identified in other non-pathogenic Xanthomonas (NPX) strains. Further investigation using multiple genomic-based taxonomic indices indicates that these strains represent a potential new species. Hence, we propose Xanthomonas imtechensis sp. nov. as a new species of the genus Xanthomonas, with the type strain being PPL568 = MTCC 13186 = CFBP 9040 = ICMP 24395.

Bacteria of the genus Achromatium are known for their large cell sizes and intracellular calcium carbonate deposits. Achromatium inhabit freshwater, brackish, and marine sediments where they accumulate to high abundances at the oxic-anoxic interface. These bacteria alter their vertical position in the sediment along with daily fluctuations in oxygen concentrations. Yet, the mechanism behind their migration in the sediment remains unknown. In this study, we used chemotaxis assays and time-lapse microphotography to analyze the motility and chemotactic behavior of Achromatium oxaliferum. Microscopic observations revealed that rolling and gliding were the main forms of locomotion exhibited by Achromatium. In absence of any stimulant, the movement appeared to be mostly random and changes in direction frequently occurred. Chemotaxis assays showed a negative chemotaxis of Achromatium to oxygen, sulfide, and nitrate, as evidenced by the change from undirected to directed locomotion against the respective chemical gradient. For periods of more than 1 hour, Achromatium cells moved continuously towards regions of low concentration. We further investigated whether the genetic repertoire of Achromatium corresponds to our observations. Based on lab experiments and bioinformatic analyses we conclude that Achomatium motility is propelled by type IV pili guided by a plethora of chemo- and photoreceptors. We conclude that Achromatium uses negative chemo- and phototaxis to confine their distribution in aquatic sediments between opposing oxygen and sulfide gradients. This allows Achromatium to dynamically adjust its position in redox gradients, and thus is likely to have a major contribution to its success in the global colonization of diverse aquatic sediments.

The genus Exiguobacterium comprises Gram-positive, non-spore-forming, facultative anaerobic bacteria known for their remarkable adaptability to extreme environments, including soils, hot springs, glaciers, and the gastrointestinal tracts of certain organisms. Despite their unique adaptations for surviving in extreme environments, their pathogenicity is well documented. Here, we analyzed the phenotypical traits of two Mexican strains of Exiguobacterium--JVH47, isolated from contaminated urban sediments in Mexico City, and P4526, from the less human-impacted Cuatro Cienegas Basin. Furthermore, strains were related via comparative genomics using publicly available genomes. Phenotypic characterization demonstrated that both strains thrive across a wide range of temperatures (20-50 {degrees}C), pH (7-11), and salinity (up to 7% NaCl). Although sensitive to erythromycin, the JVH47 strain exhibited higher erythromycin resistance and harbored antibiotic resistance genes. This study underscores the ecological versatility of Exiguobacterium and its potential role as a reservoir for antibiotic resistance genes. While rarely associated with human infections, its ability to survive in extreme conditions and form biofilms raises concerns for immunocompromised individuals. These findings highlight the need for careful consideration of Exiguobacterium in biotechnological applications and its implications under the One Health framework.

The interactions between microalgae and the bacteria living in the phycosphere are pivotal to the role they play in aquatic ecosystems. This study examines how two representatives of common phycosphere bacteria, Yoonia sp. TsM2_T14_4 (Rhodobacteraceae) and Maribacter sp. IgM3_T14_3 (Flavobacteriaceae), interact with three microalgal hosts: Isochrysis galbana, Tetraselmis suecica, and Conticribra weissflogii (formerly Thalassiosira weissflogii) using dual transcriptomic analyses of both bacteria and microalgae. Bacterial transcriptomes differed significantly depending on microalgal host, with notable changes in carbohydrate metabolism among other COG categories. Yoonia sp. expressed genes involved in anoxygenic photosynthesis in co-culture with I. galbana, presumably due to its inability to utilize carbohydrates from this algal host, whereas Maribacter sp. expressed polysaccharide degradation genes in co-culture with C. weissflogii along with T9SS genes, which can be employed to secrete these hydrolytic enzymes. Specifically, a putative glucan endo-1,3-beta-D-glucosidase was highly expressed, an enzyme that can hydrolyze laminarin and curdlan. Maribacter sp. IgM3_T14_3 could utilize laminarin as a sole carbon source in laboratory settings, a polysaccharide commonly found in marine environments and produced by C. weissflogii. Surprisingly, microalgal transcriptomes remained largely unaltered in the presence of either of the bacteria compared to transcriptomes of axenic algal cultures. These findings highlight the adaptability of phycosphere bacteria to different microalgal hosts. Furthermore, it also indicates a commensalism between microalgae, Yoonia sp. and Maribacter sp., in which the bacteria adapt to and benefit from microalgal host exudates, whereas under the conditions employed here the microalgae are unaffected by the presence of these bacterial symbionts. ImportanceMicroalgae are the key players in marine ecosystems, capturing carbon dioxide through photosynthesis and releasing carbohydrates into their immediate environment, the so-called phycosphere. Certain bacterial taxa are consistently found within the phycosphere, where they interact with their microalgal host in a variety of ways. However, the impact of these bacteria on the microalgae is not fully understood despite their ecological relevance. This study uses a dual transcriptomic approach to investigate the impact of such core phycosphere bacteria on microalgal hosts and vice versa to uncover the reason behind their success in the phycosphere and possible roles in marine ecosystems.

The strain LEGE 10371, isolated from the surface of a marine sponge at Praia da Memoria, Portugal, was characterized as a new Thalassoporum species (Pseudanabaenales) using a polyphasic approach that included 16S rRNA gene phylogenetic analysis (Maximum Likelihood and Bayesian Inference), 16S-23S ITS secondary structures, p-distance calculations, MALDI-TOF MS profiling, and morphological analysis by optical and scanning electron microscopy, as well as ecological and biochemical characterization. Phylogenetically, LEGE 10371 clustered within the Thalassoporum clade, however distant from the other existent species of the genus. The p-distance analysis revealed low sequence identity with other Thalassoporum species, with a maximum value of 97.2% to Th. komareki. The MALDI-TOF profile displayed high-intensity peaks at approximately 3,000, 4,000, 6,000 and 8,000 m/z, representing strong candidates for diagnostic markers of the new species. Morphologically, the new species differ from the other species of the genus by presenting trichomes with more than 10 cells and lack of aerotopes. Biocompatibility of the fractions was evaluated in HaCaT keratinocytes, showing no cytotoxic effects at most tested concentrations. PCR screening targeting mcyE, sxtG, anaC, and cyrA confirmed the absence of the genetic potential for the production of major cyanotoxins. Chemical characterization revealed a pigment-rich profile dominated by chlorophyll-a and carotenoids, including {beta}-carotene, zeaxanthin, lutein, and mixoxanthophyll. Bioactivity assays showed superoxide anion radical scavenging by the aqueous fraction (IC2 {approx} 0.042-0.045 mg mL-{superscript 1}), strong nitric oxide radical scavenging by the acetonic fraction (IC = 0.045 mg mL-{superscript 1}), and lipoxygenase inhibition ([~]41%, for a fraction concentration of 0.25 mg mL-), suggesting a potential contribution of these fractions to modulate inflammation-related pathways. Additionally to this results, the polyphasic analysis permitted to confirm previous data that Pseudanabaena and Limnothrix represent the same generic entity. Both genera clustered together, presented high 16S rRNA gene identity (up to 99.9%) and share the same morphological and ecological features. Consequently, we formally proposed the synonimization of Limnothrix into Pseudanabaena.

The urgent need for sustainable solutions to mitigate climate change has intensified research into carbon capture, utilization, and storage (CCUS) strategies. Biological approaches, particularly involving extremophilic microorganisms, offer promising alternatives to conventional methods due to their adaptability and potential for bioproduct synthesis. In this study, we report the complete genome sequencing and functional characterization of isolate BS253, derived from a hypersaline alkaline lake in Brazils Pantanal region. Using a hybrid sequencing strategy combining Oxford Nanopore long reads and Illumina short reads, we assembled a circular chromosome of 3.76 Mb and identified two plasmids. Phylogenetic and comparative genomic analyses identified the isolate as Vreelandella zhaodongensis. Digital DNA-DNA hybridization (dDDH % = 71.4%) and ANI (96,83%) values supported the designation of BS253 as a distinct subspecies of V. zhaodongensis. The genome reveals genes associated with salt and alkali tolerance, hydrocarbon and plastic degradation, and the biosynthesis of secondary metabolites. Phenotypically, BS253 is a moderately halophilic, facultatively anaerobic, Gram-negative rod exhibiting biosurfactant activity, with an emulsification index of 51.7% under defined culture conditions. These findings highlight BS253 as a metabolically versatile extremophile with potential applications in different types of industries and biotechnological CCUS systems. ImportanceMicroorganisms adapted to extreme environments represent an untapped source of biotechnologically valuable traits. Vreelandella zhaodongensis BS253, isolated from a hypersaline alkaline lake in the Brazilian Pantanal, expands the known diversity of extremophiles and offers metabolic features with relevance to sustainable bioprocesses. Its complete genome reveals genes involved in salt and alkali tolerance, plastic and hydrocarbon degradation, and the biosynthesis of biosurfactant-like compounds, positioning this strain as a promising chassis for applications in emerging carbon capture, utilization, and storage (CCUS) strategies. The ability of BS253 to produce bioemulsifying molecules under defined nutritional conditions, combined with pathways for degrading recalcitrant pollutants, reinforces its potential for environmentally friendly industrial processes. By characterizing BS253 at the genomic and physiological levels, this work provides foundational information for future exploitation of extremophiles in biotechnological innovations aimed at reducing carbon emissions and supporting circular bioeconomy initiatives.

Exopolysaccharides (EPS) produced by lactic acid bacteria (LAB) and other microorganisms have attracted considerable interest due to their structural diversity and physicochemical properties, which makes them valuable across various industrial applications. To achieve high cell densities and maximize EPS yields, microorganisms are typically cultivated in nutrient-rich media containing yeast extract. However, yeast extract may contain high molecular weight polysaccharides that are not metabolized by the bacteria. This can lead to an overestimation of EPS yields and contamination of the bacterial EPS, potentially resulting in misinterpretation of their structure and biological activity. In this study, we demonstrate the presence of high molecular weight -mannan and {beta}-glucan in yeast extract in EPS isolates using both ultrafiltration and the commonly used trichloroacetic acid/ethanol (TCA/EtOH) precipitation method. These polysaccharides were characterized by size-exclusion chromatography, high-performance anion-exchange chromatography, and nuclear magnetic resonance spectroscopy. Their abundances were estimated to range from 10 to 50 mg/L in MRS medium, depending on the supplier of the yeast extract. The main contaminant identified was yeast -mannan. By cultivating L. rhamnosus GG (ATCC 53103) and L. pentosus KW1 and isolating their respective EPS, we illustrate how these yeast extract contaminants affect the structural interpretation of the EPS and that the contaminants can be completely removed by ultrafiltration of the growth medium prior to bacterial cultivation. In conclusion, we emphasize the necessity of stringent controls in the production and purification of microbial EPS, with particular attention to the chemical purity of medium constituents.

The Hawaiian islands are among the most geologically and volcanically active places on Earth. While the Hawaiian Archipelago is known for its animal and plant diversity, much less is known about microbial diversity in the areas diverse habitats. In this study, we focused on steam vent associated biofilms found on the most volcanically active island of Hawai{square}i, also known as the Big Island. From 46 samples from various biofilms and associated features around fumaroles emitting water steam, we generated amplicon and metagenomic sequences. Amplicon data showed that Chloroflexota and Cyanobacteriota are the numerically dominant phyla in these biofilm communities. We constructed 363 non-redundant medium to high-quality metagenome-assembled genomes (MAGs) that are at least 70% complete and with less than 5% contamination. Ten MAGs belong in the domain Archaea, and 353 belong in the domain Bacteria. This dataset could provide valuable insights into microbial diversity and ecology around volcanic features in Hawai i and elsewhere.

Mixotrophy is emerging as a default nutritional strategy in phytoplankton but research seems so far isolated and mostly focussing on single phytoplankton groups or strains. Here we combined data from 24 oceanic and 22 freshwater strains - as well as results from other studies - to analyze phytoplanktons ability to utilize dissolved organic compounds and highlight potential influencing factors. The results emphasize that mixotrophy is ubiquitous in phytoplankton across functional groups and taxa isolated from various habitats, and not strictly dependent on light or nutrient deficiencies. Several factors such as taxonomic affiliation, temperature and growth phase can affect mixotrophic behavior but no consistent patterns have emerged regarding their effects. Hence, mixotrophic traits remain so far unpredictable. There is some indication that the strains origin - potentially through adaptation to habitat DOM availability - might predetermine phytoplanktons mixotrophic skills. For example, freshwater strains used overall more compounds than oceanic strains in our study and Ostreococcus exhibited a different use pattern depending on its origin. Nevertheless, many aspects of mixotrophy in phytoplankton - e.g. metabolic pathways - remain cryptic. By summarizing available knowledge and knowledge gaps, the present synthesis provides a guideline for upcoming research further exploring mixotrophy. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=135 SRC="FIGDIR/small/703725v1_ufig1.gif" ALT="Figure 1"> View larger version (47K): org.highwire.dtl.DTLVardef@12ac311org.highwire.dtl.DTLVardef@6c98deorg.highwire.dtl.DTLVardef@1a837ddorg.highwire.dtl.DTLVardef@eb9b53_HPS_FORMAT_FIGEXP M_FIG C_FIG

The microbiota and microbiome associated with zooplankton remains rather understudied compared to other animal groups and/or taxa. The present study aimed at investigating the whole-body bacterial microbiota of Daphnia spp. in two contrasting Greek lakes, the shallow and hypertrophic Lake Koronia vs. the deep and mesotrophic Lake Vegoritida, including both egg-bearing and non-egg-bearing individuals. In both lakes, 2,060 bacterial operational taxonomic units (OTUs) were found, with 223 of them being conditionally rare (crOTUs) with low contribution even for the dominant phyla, with L. Vegoritida having more crOTUs than L. Koronia. The individuals microbiota had inconsiderable overlap with the surrounding water microbiota in both lakes. The two lakes showed significant differences in their Daphnia -associated microbiota. L. Koronia had richer OTUs and rather homogeneous bacterial communities, with higher occupancy. Overall, no significant differences in between the microbiota of egg-bearing and non-egg-bearing Daphnia individuals in both lakes. However, regarding the most important OTUs (miOTUs), the L. Koronia miOTUs were highly overlapped between the individuals with and without eggs, with only one missing from the individuals without eggs. In L. Vegoritida the individuals without eggs had only six miOTUs and while egg-bearing individuals had nine different ones; the two lakes had no shared miOTUs., considerable differences occurred.. A total of 27 miOTUs, was found and belonged to the Pseudomonadota, unclassified Bacteria, Cyanobacteria, Bacteroidota, Bacillota and Actinomycetota. Those miOTUs, where assignment to the genus level was possible, they were related to Cyanobium, Mucilaginibacter, Flavobacterium and Staphylococcus. This study showed that lake morphotype and ecological status can exert some impact on Daphnia-associated bacterial microbiota, with more pronounced effects on egg-bearing and non-egg-bearing individuals.

Prokaryotes, particularly those in extreme environments, are capable of diverse metabolic states resulting in altered cell envelope structure and function. However, these changes are difficult to assess as standard fluorescent probes are often incompatible with extreme conditions and/or extremophile cell physiology. Halophilic archaea present the challenge of near-saturated intra-/extra-cellular salts, high membrane potential, and extended survival in altered metabolic states including entrapped within salt crystal fluid inclusions. We evaluated the compatibility of six fluorescent markers of cell envelope stability and activity with two model species, Halobacterium salinarum and Haloferax volcanii. Redox activity markers alamarBlue and pure resazurin solutions, membrane potential probes MitoTracker Orange-CMTMRos and Rhodamine 123, and SYTO 9 and propidium iodide (LIVE/DEAD kit) to assess cell membrane integrity were evaluated for use in bulk (microplate reader) and cell-specific (microscopy) applications. Limitations of each probe were identified, clarifying the utilization of each based on cell physiology, growth phase, medium composition, and probe exposure time including extended timescales needed to simulate the environmental conditions of haloarchaea. Of particular note, propidium iodide behavior was unreliable leading to double-labeling of cells and false interpretation of cells as dead. These data provide important insights into the study of prokaryotes in non-standard conditions.