Estimation of baseline levels of bacterial community tolerance to Cr, Ni, Pb, and Zn in unpolluted soils, a background for PICT (pollution-induced community tolerance) determination

The PICT method (pollution-induced community tolerance) can be used to assess whether changes in soil microbial response are due to heavy metal toxicity or not. Microbial community tolerance baseline levels can, however, also change due to variations in soil physicochemical properties. Thirty soil samples (0–20 cm), with geochemical baseline concentrations (GBCs) of heavy metals and from five different parent materials (granite, limestone, schist, amphibolite, and serpentine), were used to estimate baseline levels of bacterial community tolerance to Cr, Ni, Pb, and Zn using the leucine incorporation method. General equations (n = 30) were determined by multiple linear regression using general soil properties and parent material as binary variables, explaining 38% of the variance in log IC50 (concentration that inhibits 50% of bacterial growth) values for Zn, with 36% for Pb, 44% for Cr, and 68% for Ni. The use of individual equations for each parent material increased the explained variance for all heavy metals, but the presence of a low number of samples (n = 6) lead to low robustness. Generally, clay content and dissolved organic C (DOC) were the main variables explaining bacterial community tolerance for the tested heavy metals. Our results suggest that these equations may permit applying the PICT method with Zn and Pb when there are no reference soils, while more data are needed before using this concept for Ni and Cr.

Bacterial Abundance, Diversity and Activity During Long-Term Colonization of Non-biodegradable and Biodegradable Plastics in Seawater

The microorganisms living on plastics called “plastisphere” have been classically described as very abundant, highly diverse, and very specific when compared to the surrounding environments, but their potential ability to biodegrade various plastic types in natural conditions have been poorly investigated. Here, we follow the successive phases of biofilm development and maturation after long-term immersion in seawater (7 months) on conventional [fossil-based polyethylene (PE) and polystyrene (PS)] and biodegradable plastics [biobased polylactic acid (PLA) and polyhydroxybutyrate-co-hydroxyvalerate (PHBV), or fossil-based polycaprolactone (PCL)], as well as on artificially aged or non-aged PE without or with prooxidant additives [oxobiodegradable (OXO)]. First, we confirmed that the classical primo-colonization and growth phases of the biofilms that occurred during the first 10 days of immersion in seawater were more or less independent of the plastic type. After only 1 month, we found congruent signs of biodegradation for some bio-based and also fossil-based materials. A continuous growth of the biofilm during the 7 months of observation (measured by epifluorescence microscopy and flow cytometry) was found on PHBV, PCL, and artificially aged OXO, together with a continuous increase in intracellular (3H-leucine incorporation) and extracellular activities (lipase, aminopeptidase, and β-glucosidase) as well as subsequent changes in biofilm diversity that became specific to each polymer type (16S rRNA metabarcoding). No sign of biodegradation was visible for PE, PS, and PLA under our experimental conditions. We also provide a list of operational taxonomic units (OTUs) potentially involved in the biodegradation of these polymers under natural seawater conditions, such as Pseudohongiella sp. and Marinobacter sp. on PCL, Marinicella litoralis and Celeribacter sp. on PHBV, or Myxococcales on artificially aged OXO. This study opens new routes for a deeper understanding of the polymers’ biodegradability in seawaters, especially when considering an alternative to conventional fossil-based plastics.

Changes in microbial community phylogeny and metabolic activity along the water column uncouple at near sediment aphotic layers in fjords

Fjords are semi-enclosed marine systems with unique physical conditions that influence microbial community composition and structure. Pronounced organic matter and physical condition gradients within fjords provide a natural laboratory for the study of changes in microbial community structure and metabolic potential in response to environmental conditions. Photosynthetic production in euphotic zones sustains deeper aphotic microbial activity via organic matter sinking, augmented by large terrestrial inputs. Previous studies do not consider both prokaryotic and eukaryotic communities when linking metabolic potential and activity, community composition, and environmental gradients. To address this gap we profiled microbial functional potential (Biolog Ecoplates), bacterial abundance, heterotrophic production (3H-Leucine incorporation), and prokaryotic/eukaryotic community composition (16S and 18S rRNA amplicon gene sequencing). Similar factors shaped metabolic potential, activity and community (prokaryotic and eukaryotic) composition across surface/near surface sites. However, increased metabolic diversity at near bottom (aphotic) sites reflected an organic matter influence from sediments. Photosynthetically produced particulate organic matter shaped the upper water column community composition and metabolic potential. In contrast, microbial activity at deeper aphotic waters were strongly influenced by other organic matter input than sinking marine snow (e.g. sediment resuspension of benthic organic matter, remineralisation of terrestrially derived organic matter, etc.), severing the link between community structure and metabolic potential. Taken together, different organic matter sources shape microbial activity, but not community composition, in New Zealand fjords.

Tolerance of soil bacterial community to tetracycline antibiotics induced by As, Cd, Zn, Cu, Ni, Cr and Pb pollution

The widespread use of both heavy metals and antibiotics in livestock farming and their subsequent arrival on agricultural soils through manure/slurry spreading has become a problem of vital importance for human health and the environment. In the current research, a laboratory experiment was carried out for 42 days to study co-selection for tolerance of three tetracycline antibiotics (tetracycline, TC; oxytetracycline, OTC; chlortetracycline, CTC) in soils polluted with heavy metals (As, Cd, Zn, Cu, Ni, Cr and Pb) at high concentration levels (1000 mg kg−1 of each one, separately). Pollution Induced Community Tolerance (PICT) of the bacterial community was estimated using the leucine incorporation technique. The Log IC50 (logarithm of the concentration causing 50 % inhibition in bacterial community growth) values obtained in uncontaminated soil samples for all the heavy metals tested showed the following toxicity sequence: Cu > As > Cr ≥ Pb ≥ Cd > Zn > Ni. However, in polluted soil samples the toxicity sequence was: Cu > Pb ≥ As ≥ Cd ≥ Cr ≥ Ni ≥ Zn. Moreover, at high metal concentrations the bacterial communities show tolerance to the metal itself, this taking place for all the metals tested in the long term. The bacterial communities of the soil polluted with heavy metals showed also long-term co-tolerance to TC, OTC, and CTC. This kind of studies, focusing on the eventual increases of tolerance and co-tolerance of bacterial communities in agricultural soil, favored by the presence of other pollutants, is of crucial importance, mostly bearing in mind that the appearance of antibiotic resistance genes in soil bacteria could be transmitted to human pathogens.

Changes in Microbial Community Phylogeny and Metabolic Activity Along the Water Column Uncouple at Near Sediment Aphotic Layers in Fjords

Fjords are semi-enclosed marine systems with unique physical conditions that influence microbial community composition and structure. Pronounced organic matter and physical condition gradients within fjords provide a natural laboratory for the study of changes in microbial phylogeny and metabolic potential in response to environmental conditions. Photosynthetic production in euphotic zones sustains deeper aphotic microbial activity via organic matter sinking, augmented by large terrestrial inputs. We profiled microbial functional potential (Biolog Ecoplates), bacterial abundance, heterotrophic production (3H-Leucine incorporation), and prokaryotic/eukaryotic community composition (16S and 18S rRNA amplicon gene sequencing) to link metabolic potential, activity, and community composition to known community drivers. Similar factors shaped metabolic potential, activity and community (prokaryotic and eukaryotic) composition across surface/near surface sites. However, increased metabolic diversity at near bottom (aphotic) sites reflected an organic matter influence from sediments. Photosynthetically produced particulate organic matter shaped the upper water column community composition and metabolic potential. In contrast, microbial activity at deeper aphotic waters were strongly influenced by other organic matter imput than sinking marine snow (e.g. sediment resuspension of benthic organic matter, remineralisation of terrestrially derived organic matter, etc.), severing the link between phylogeny and metabolic potential. Taken together, different organic matter sources shape microbial activity, but not community composition, in New Zealand fjords.

Relative Influence of Plastic Debris Size and Shape, Chemical Composition and Phytoplankton-Bacteria Interactions in Driving Seawater Plastisphere Abundance, Diversity and Activity

The thin film of life that inhabits all plastics in the oceans, so-called “plastisphere,” has multiple effects on the fate and impacts of plastic in the marine environment. Here, we aimed to evaluate the relative influence of the plastic size, shape, chemical composition, and environmental changes such as a phytoplankton bloom in shaping the plastisphere abundance, diversity and activity. Polyethylene (PE) and polylactide acid (PLA) together with glass controls in the forms of meso-debris (18 mm diameter) and large-microplastics (LMP; 3 mm diameter), as well as small-microplastics (SMP) of 100 μm diameter with spherical or irregular shapes were immerged in seawater during 2 months. Results of bacterial abundance (confocal microscopy) and diversity (16S rRNA Illumina sequencing) indicated that the three classical biofilm colonization phases (primo-colonization after 3 days; growing phase after 10 days; maturation phase after 30 days) were not influenced by the size and the shape of the materials, even when a diatom bloom (Pseudo-nitzschia sp.) occurred after the first month of incubation. However, plastic size and shape had an effect on bacterial activity (3H leucine incorporation). Bacterial communities associated with the material of 100 μm size fraction showed the highest activity compared to all other material sizes. A mature biofilm developed within 30 days on all material types, with higher bacterial abundance on the plastics compared to glass, and distinct bacterial assemblages were detected on each material type. The diatom bloom event had a great impact on the plastisphere of all materials, resulting in a drastic change in diversity and activity. Our results showed that the plastic size and shape had relatively low influence on the plastisphere abundance, diversity, and activity, as compared to the plastic composition or the presence of a phytoplankton bloom.

Changes in Activity and Community Composition Shape Bacterial Responses to Size-Fractionated Marine DOM

To study the response of bacteria to different size-fractions of naturally occurring dissolved organic matter (DOM), a natural prokaryotic community from North Atlantic mesopelagic waters (1000 m depth) was isolated and grown in (i) 0.1-μm filtered seawater (CONTROL), (ii) the low-molecular-weight (<1 kDa) DOM fraction (L-DOM), and (iii) the recombination of high- (>1 kDa) and low-molecular-weight DOM fractions (H + L-DOM), to test the potential effect of ultrafiltration on breaking the DOM size continuum. Prokaryotic abundance and leucine incorporation were consistently higher in the H + L-DOM niche than in the L-DOM and CONTROL treatments, suggesting a different interaction with each DOM fraction and the disruption of the structural DOM continuum by ultrafiltration, respectively. Rhodobacterales (Alphaproteobacteria) and Flavobacteriales (Bacteroidetes) were particularly enriched in L-DOM and closely related to the colored DOM (CDOM) fraction, indicating the tight link between these groups and changes in DOM aromaticity. Conversely, some other taxa that were rare or undetectable in the original bacterial community were enriched in the H + L-DOM treatment (e.g., Alteromonadales belonging to Gammaproteobacteria), highlighting the role of the rare biosphere as a seed bank of diversity against ecosystem disturbance. The relationship between the fluorescence of protein-like CDOM and community composition of populations in the H + L-DOM treatment suggested their preference for labile DOM. Conversely, the communities growing on the L-DOM niche were coupled to humic-like CDOM, which may indicate their ability to degrade more reworked DOM and/or the generation of refractory substrates (as by-products of the respiration processes). Most importantly, L- and/or H + L-DOM treatments stimulated the growth of unique bacterial amplicon sequence variants (ASVs), suggesting the potential of environmental selection (i.e., changes in DOM composition and availability), particularly in the light of climate change scenarios. Taken together, our results suggest that different size-fractions of DOM induced niche-specialization and differentiation of mesopelagic bacterial communities.

Effect of Oxytetracycline and Chlortetracycline on Bacterial Community Growth in Agricultural Soils

Toxicity on soil bacterial community growth caused by the antibiotics oxytetracycline (OTC) and chlortetracycline (CTC) was studied in 22 agricultural soils after 1, 8 and 42 incubation days. The leucine incorporation method was used with this aim, estimating the concentration of each antibiotic which caused an inhibition of 50% in bacterial community growth (log IC50). For OTC, the mean log IC50 was 2.70, 2.81, 2.84 for each of the three incubation times, while the values were 2.05, 2.22 and 2.47 for CTC, meaning that the magnitude of OTC toxicity was similar over time, whereas it decreased significantly for CTC with incubation time. In addition, results showed that the toxicity on bacterial community growth due to CTC is significantly higher than when due to OTC. Moreover, the toxicity on bacterial community growth due to both antibiotics is dependent on soil properties. Specifically, an increase in soil pH and silt content resulted in higher toxicity of both antibiotics, while increases in total organic carbon and clay contents caused decreases in OTC and CTC toxicities. The results also show that OTC toxicity can be well predicted by means of specific equations, using the values of pH measured in KCl and those of effective cation exchange capacity as input variables. CTC toxicity may be predicted (but with low precision) using pH measured in KCl and total organic carbon. These equations may help to predict the negative effects caused by OTC and CTC on soil bacteria using easily measurable soil parameters.

Molecular Mechanisms of Adiponectin-Induced Attenuation of Mechanical Stretch-Mediated Vascular Remodeling

Hypertension induces vascular hypertrophy, which changes blood vessels structurally and functionally, leading to reduced tissue perfusion and further hypertension. It is also associated with dysregulated levels of the circulating adipokines leptin and adiponectin (APN). Leptin is an obesity-associated hormone that promotes vascular smooth muscle cell (VSMC) hypertrophy. APN is a cardioprotective hormone that has been shown to attenuate hypertrophic cardiomyopathy. In this study, we investigated the molecular mechanisms of hypertension-induced VSMC remodeling and the involvement of leptin and APN in this process. To mimic hypertension, the rat portal vein (RPV) was mechanically stretched, and the protective effects of APN on mechanical stretch-induced vascular remodeling and the molecular mechanisms involved were examined by using 10 μg/ml APN. Mechanically stretching the RPV significantly decreased APN protein expression after 24 hours and APN mRNA expression in a time-dependent manner in VSMCs. The mRNA expression of the APN receptors AdipoR1, AdipoR2, and T-cadherin significantly increased after 15 hours of stretch. The ratio of APN/leptin expression in VSMCs significantly decreased after 24 hours of mechanical stretch. Stretching the RPV for 3 days increased the weight and [3H]-leucine incorporation significantly, whereas APN significantly reduced hypertrophy in mechanically stretched vessels. Stretching the RPV for 10 minutes significantly decreased phosphorylation of LKB1, AMPK, and eNOS, while APN significantly increased p-LKB1, p-AMPK, and p-eNOS in stretched vessels. Mechanical stretch significantly increased p-ERK1/2 after 10 minutes, whereas APN significantly reduced stretch-induced ERK1/2 phosphorylation. Stretching the RPV also significantly increased ROS generation after 1 hour, whereas APN significantly decreased mechanical stretch-induced ROS production. Exogenous leptin (3.1 nM) markedly increased GATA-4 nuclear translocation in VSMCs, whereas APN significantly attenuated leptin-induced GATA-4 nuclear translocation. Our results decipher molecular mechanisms of APN-induced attenuation of mechanical stretch-mediated vascular hypertrophy, with the promising potential of ultimately translating this protective hormone into the clinic.