Exploring Rhodospirillum rubrum response to high doses of carbon monoxide under light and dark conditions.

Environmental concerns about residues and the traditional disposal methods are driving the search for more environmentally conscious processes, such as pyrolysis and gasification. Their main final product is synthesis gas (syngas) composed of CO, CO2, H2, and methane. Syngas can be converted into various products using CO-tolerant microorganisms. Among them, Rhodospirillum rubrum is highlighted for its biotechnological potential. However, the extent to which high doses of CO affect its physiology is still opaque. For this reason, we have studied R. rubrum behavior under high levels of this gas (up to 2.5 bar), revealing a profound dependence on the presence or absence of light. In darkness, the key variable affected was the lag phase, where the highest levels of CO retarded growth to more than 20 days. Under light, R. rubrum ability to convert CO into CO2 and H2 depended on the presence of an additional carbon source, such as acetate. In those conditions where CO was completely exhausted, CO2 fixation was unblocked, leading to a diauxic growth. To enhance R. rubrum tolerance to CO in darkness, a UV-accelerated adaptive laboratory evolution (UVa-ALE) trial was conducted to isolate clones with shorter lag phases, resulting in the isolation of clones 1.4-2B and 1.7-2A. The adaptation of 1.4-2B was mainly based on mutated enzymes with a metabolic function, while 1.7-3A was mostly affected at regulatory genes, including the anti-repressor PpaA/AerR. Despite these mutations having slight effects on biomass and pigment levels, they successfully provoked a significant reduction in the lag phase (-50%). KEYPOINTS: • CO affects principally R. rubrum lag phase (darkness) and growth rate (light) • CO is converted to CO2/H2 during acetate uptake and inhibits CO2 fixation (light) • UVa-ALE clones showed a 50% reduction in the lag phase (darkness).

Leaching of herbicides mixtures from pre-exposed agricultural plastics severely impact microalgae.

Farmlands represent a source of aged plastics and pesticides to the surrounding environments. It has been shown that chemicals can be sorbed and desorbed from plastics, but the interaction between plastic and mixtures of pesticides and their effects on freshwater biota has not been assessed yet. The aim of the work was to assess the potential role of agricultural plastics as vectors for a mixture of two herbicides and the impact of the herbicide mixture lixiviated from them towards the freshwater microalga Chlamydomonas reinhardtii. Pristine and aged polyethylene plastics collected from agricultural areas were exposed to the herbicides, bifenox, oxyfluorfen and their mixtures. The microalgae were exposed for 72 h to the leachates desorbed from plastics and the effect was quantified in terms of total chlorophyll content and several physiological parameters assessed by flow cytometry. Our results showed that changes in physicochemical properties (hydroxyl and carbonyl index, hydrophobicity, texture) in aged plastics increased their capacity to retain and to desorb the herbicides. Microalgae exposed to leachates containing bifenox, oxyfluorfen, or their mixture showed reactive oxygen species overproduction, lipid peroxidation, membrane potential hyperpolarization, intracellular pH acidification, and a loss of metabolic activity. The toxicological interactions of the leachate mixture were assessed using the Combination Index (CI)-isobologram method showing antagonism at low effect levels turning to synergism when the effect increased. In this work, we proved the hypothesis that ageing increases the capacity of agricultural plastics to behave as vector for toxic chemicals to the biota.

Biofilm formation strongly influences the vector transport of triclosan-loaded polyethylene microplastics.

This study aimed at evaluating the influence of biofilm in the role of microplastics (MPs) as vectors of pollutants and their impact on Daphnia magna. To do this, virgin polyethylene MPs, (PE-MPs, 40-48 µm) were exposed for four weeks to wastewater (WW) from influent and effluent to promote biofouling. Then, the exposed PE-MPs were put in contact with triclosan. Finally, the toxicity of TCS-loaded and non-TCS loaded PE-MPs were tested on the survival of D. magna adults for 21 days. Results from metabarcoding analyses indicated that exposure to TCS induced shifts in the bacterial community, selecting potential TCS-degrading bacteria. Results also showed that PE-MPs were ingested by daphnids. The most toxic virgin PE-MPs were those biofouled in the WW effluent. The toxicity of TCS-loaded PE-MPs biofouled in the WW effluent was even higher, reporting mortality in all tested concentrations. These results indicate that biofouling of MPs may modulate the adsorption and subsequent desorption of co-occurring pollutants, hence affecting their potential toxicity towards aquatic organisms. Future studies on realistic environmental plastic impact should include the characterization of biofilms growing on plastic. Since inevitably plastic biofouling occurs over time in nature, it should be taken into account as it may modulate the sorption of co-occurring pollutants.

Polystyrene nanoplastics and wastewater displayed antagonistic toxic effects due to the sorption of wastewater micropollutants.

The knowledge about the interaction of nanoplastics with other aquatic pollutants and their combined effects on biota is very scarce. In this work, we studied the interaction between polystyrene nanoplastics (PS NPs) (30 nm) and the micropollutants in a biologically treated wastewater effluent (WW). The capacity of PS NPs to sorb micropollutants was studied as well as their single and combined toxicity towards three freshwater organisms: the recombinant bioluminescent cyanobacterium, Anabaena sp. PCC 7120 CPB4337; the duckweed, Spirodela polyrhiza and the cladoceran, Daphnia magna. The endpoints were the inhibition of bioluminescence, the growth inhibition of the aquatic plant and the immobilization of D. magna after 24, 72 and 48 h of exposure, respectively. Combination Index (CI)-isobologram method was used to quantify mixture toxicity and the nature of interactions. PS NPs sorbed a variety of chemicals present in WW as micropollutants in a range of tens of ng/L to µg/L. It was found that those pollutants with positive charge were the main ones retained onto PS NPs, which was attributed to the electrostatic interaction with the negatively charged PS NPs. Regarding the toxicological effects, single exposure to PS NPs affected the three tested organisms. However, single exposure to WW only had a negative impact on the cyanobacterium and S. polyrhiza with no observed toxicity to D. magna. Regarding PS NPs-WW combined exposure, a reduction of toxicity in comparison with single exposure was observed probably due to the sorption of micropollutants onto PS NPs, which resulted in lower bioavailability of the micropollutants. In addition, the formation of PS NPs-WW heteroaggregates was observed which could result in lower bioavailability of PS NPs and sorbed micropollutants, thus lowering toxicity. This study represents a near-realistic scenario approach to the potential sorption of wastewater pollutants onto nanoplastics that could alter the toxicological effect on the biota.

Microplastics can act as vector of the biocide triclosan exerting damage to freshwater microalgae.

Despite the large number of recent studies on microplastics (MPs) and their ability to act as carriers of pollutants, the knowledge about the biological effects of MPs loaded with chemicals is scarce. The aim of this study was to evaluate the potential of MPs as vectors for the antimicrobial triclosan (TCS). For it, we tested low-density polyethylene (LDPE), polyamide (PA), polyethylene terephthalate (PET), polyoxymethylene (POM), polypropylene (PP), polystyrene (PS) and the biodegradable polylactic acid (PLA). Thus, chemical analysis of sorption and desorption of TCS by these MPs was evaluated. The effect of TCS-loaded MPs to Anabaena sp. PCC7120, a cyanobacterium model of primary producers in freshwater ecosystems, was investigated. Chemical analyses showed different capacity of sorption depending on the MP type, which was related to some of their physicochemical properties. PA (104.7 µg/g), POM (57.4 µg/g) and LDPE (18.3 µg/g) were the polymers that sorbed the highest amounts of TCS. Glass transition temperature of polymers and their physicochemical interaction with TCS explained the extent of sorption. Significant decreases were found in growth, 22.3%, 94.6% and 81.0%, and chlorophyll a content, 58.4%, 95.0% and 89.6%, of Anabaena when exposed to TCS-loaded LDPE, PA and POM beads, respectively, which were the only MPs displaying significant sorption-desorption of TCS, implying that these MPs could act as vectors of TCS towards freshwater microalgae. This finding is of fundamental relevance as microalgae are at the base of the aquatic trophic chain and support growth of upper organisms.

Microplastics as vectors of the antibiotics azithromycin and clarithromycin: Effects towards freshwater microalgae.

Water pollution due to microplastics (MPs) is recognized as a major anthropogenic impact. Once MPs reach the ecosystems, they are exposed to a variety of other pollutants, which can be sorbed on them, transported and eventually desorbed. In this work, we tested the hypothesis that MPs can behave as conveyors for delivering chemicals toxic to aquatic microorganisms by investigating the vector role of MPs of polyethylene terephthalate (PET), polylactic acid (PLA), polyoxymethylene (POM) and polystyrene (PS) to the macrolide antibiotics azithromycin (AZI) and clarithromycin (CLA). AZI and CLA were chosen, as they are included in the Watch List for EU monitoring concerning water policy by Decision (EU) 2018/840. MPs were loaded in contact with 500 µg/L of AZI or 1000 µg/L of CLA. Results showed that both antibiotics were sorbed on all tested MPs. The more hydrophobic AZI was sorbed in higher proportion than CLA. Both antibiotics were desorbed from MPs upon contact with water with percentages between 14.6 ± 2.6% for AZI and 1.9 ± 1.4% for CLA of the concentrations to which the MPs were initially exposed. Virgin MPs were not toxic to the cyanobacterium Anabaena sp. PCC7120. However, antibiotic-loaded MPs significantly inhibited the growth and chlorophyll content of the cyanobacterium. Most of the sorbed antibiotics became released upon contact with cyanobacterial cultures, which was the cause for the observed toxicity. Therefore, MPs can play a role as vectors of antibiotics in freshwaters systems affecting the basic trophic level of photosynthetic microorganisms.