Biofilm enhances the interactive effects of microplastics and oxytetracycline on zebrafish intestine.

The enhanced adsorption of pollutants on biofilm-developed microplastics has been proved in many studies, but the ecotoxicological effects of biofilm-developed microplastics on organisms are still unclear. In this study, adult zebrafish were exposed to original microplastics, biofilm-developed microplastics, original microplastics absorbed with oxytetracycline (OTC), and biofilm-developed microplastics absorbed with OTC for 30 days. The intestinal histological damage, intestinal biomarker response, gut microbiome and antibiotic resistance genes (ARGs) profile of zebrafish were measured to explore the roles of biofilm in the effects of microplastics. The results showed that biofilm-developed microplastics significantly increased the number of goblet cells in intestinal epithelium compared with the control group. The biofilm-developed microplastics also induced the oxidative response in the zebrafish intestines, and biofilm changed the response mode in the combined treatment with OTC. Additionally, the biofilm-developed microplastics caused intestinal microbiome dysbiosis, and induced the abundance of some pathogenic genera increasing by several times compared with the control group and the original microplastics treatments, regardless of OTC adsorption. Furthermore, the abundance of ARGs in biofilm-developed microplastics increased significantly compared with the control and the original microplastic treatments. This study emphasized the significant influence and unique role of biofilm in microplastic studies.

Effects of biodegradable microplastics on arsenic migration and transformation in paddy soils: a comparative analysis with conventional microplastics.

The combined pollution of microplastics (MPs) and arsenic (As) in paddy soils has attracted more attention worldwide. However, there are few comparative studies on the effects of biodegradable and conventional MPs on As migration and transformation. Therefore, conventional (polystyrene, polyethylene, polyvinyl chloride) and biodegradable (polybutadiene styrene, polylactic acid, polybutylene adipate terephthalate) MPs were selected to explore and demonstrate their influences and mechanism on As migration from paddy soils to overlying water and As speciation transformation through microcosmic experiment with measuring the changes of As chemical distribution, physicochemical indexes and microbial community in paddy soils. The results showed that biodegradable MPs enhanced As migration and transformation more effective than conventional MPs during 60 d. Biodegradable MPs indirectly increased the content of As(Ⅲ) and bioavailable As by changing the microbial community structure and affecting the biogeochemical cycles of carbon, nitrogen, sulfur and iron in soils, and promoted the As migration and transformation. PBS showed the strongest promoting effect, transforming to more As(Ⅲ) (11.43%) and bioavailable As (4.28%) than control. This helps to a better understanding of the effects of MPs on As biogeochemical cycle and to clarify the ecological and food safety risks of their combined pollution in soils.

Influence of extracellular polymeric substances on arsenic bioaccumulation and biotransformation in biofilms.

It is well recognized that biofilms can biosorb and biotransform heavy metals in aquatic environments. However, the effects of extracellular polymeric substance (EPS) on inorganic arsenic (As) bioaccumulation and biotransformation in biofilms are still unrevealed and need to be investigated. In order to explore the above scientific issues, the As accumulation and speciation in EPS-containing or EPS-free biofilms and growth medium under As(V)/As(III) exposure conditions were measured. After the removal of EPS, the amount of As uptake (Asup) and As adsorption (Asad) in biofilms were significantly reduced, no matter whether exposed to As(V) or As(III). FTIR analysis further suggested that the interaction between these functional groups with As was limited after the removal of EPS. In the EPS-containing biofilms, the Asad was mainly As(V) with low toxicity. However, after the removal of EPS, the Asad was mainly As(III) with high fluidity, and no methylated As was found. Moreover, the removal of EPS inhibited As(III) oxidation and methylation by biofilms, resulting in the decrease of As(V) and methylated As in the growth medium. The findings of this study emphasized the essential impact of EPS on the biosorption and biotransformation of As in biofilms. This study provides a unique understanding of the role of biofilms in As biogeochemical cycle, and water quality purification function in water environments.

Accumulation and ecotoxicological effects induced by combined exposure of different sized polyethylene microplastics and oxytetracycline in zebrafish.

Microplastics have been widely reported as carriers of antibiotics, yet studies investigating the combined ecotoxicology of microplastics and antibiotics on organisms is limited. In this study, different sized polystyrene plastics and oxytetracycline (OTC) were used to carry out a 30-day single and binary-combined exposure experiment of zebrafish, and the microplastics and OTC accumulation, liver histological alteration, biomarkers and transcriptomic response of zebrafish were evaluated. Our results indicated that 300 nm and 50 nm plastic particles increased the OTC accumulation in liver by 33.8% and 44.5%, respectively. Microplastics and OTC induced severe liver histological damage, and the damage is size-dependent, increasing with the decrease of microplastics sizes. The liver biomarkers indicated a different response pattern in single microplastics exposure and combined with OTC, single or co-exposure of 50 nm nano-plastics and OTC induced intense responses of integrated biomarker response values. The 50 nm nano-plastics, OTC and their combined exposure induced 1330, 2693 and 3965 significantly differentially expressed genes, respectively, in which the steroid biosynthesis pathway was significantly affected by all the three treatments. This study elucidated the size-dependent effects of microplastics and provided detailed data from histopathology to transcriptome profile, enhancing our understanding of the ecotoxicity of microplastics and OTC.

Fusibacter ferrireducens sp. nov., an anaerobic, Fe(â ¢)- and sulphur-reducing bacterium isolated from mangrove sediment.

An anaerobic, alkaliphilic, halotolerant, Gram-stain-positive and rod-shaped bacterium, designated Q10-2T, was isolated from mangrove sediment sampled at the Jiulong river estuary, PR China. The cells of strain Q10-2T were motile and 0.5×2-4 µm in size. Strain Q10-2T grew at 8-45 °C (optimum, 32 °C), at pH 7.0-10.5 (optimum, pH 8.5) and in the presence of 0-6 % (w/v) NaCl (optimum, 3 %). It could use complex organic compounds and carbohydrates including d-fructose, d-galactose, d-glucose, d-mannitol, d-xylose, trehalose, lactose, maltose, sucrose and starch as carbon sources and electron donors. It could reduce sulphate, thiosulphate and elemental sulphur to sulphide, but not sulphite. Fe (Ⅲ) citrate, ferrihydrite, haematite and goethite in the presence of glucose as the electron donor were also reduced. Acetate, butyrate, ethanol, CO2 and H2 were end products of glucose fermentation. The predominant cellular fatty acids were composed of C14 : 0, C16 : 0 and summed features containing C16 : 1 ω7c and/or iso-C15 : 0 2-OH and iso-C17 : 1 and/or anteiso-C17 : 1 B. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the novel strain was most closely related to Fusibacter paucivorans DSM 12116T (95.5 % sequence similarity). The genome size of strain Q10-2T was 5.0 Mb, with a G+C content of 37.4 mol%. The average nucleotide identity and digital DNA-DNA hybridization values between strain Q10-2T and F. paucivorans DSM 12116T were 69.1 and 21.8 %, respectively. The combined genotypic and phenotypic data showed that strain Q10-2T represents a novel species of the genus Fusibacter, for which the name Fusibacter ferrireducens sp. nov. is proposed. The type strain is Q10-2T (=MCCC 1A16257T=KCTC 15906T).

Complete genome sequence of Anoxybacter fermentans DY22613T, a piezophilic dissimilatory Fe(III)-reducing bacterium isolated from East Pacific Rise hydrothermal sulfides.

Anoxybacter fermentans DY22613T is a novel piezophilic dissimilatory iron(III)-reducing bacterium isolated from East Pacific Rise (EPR) hydrothermal sulfides. The strain shows very low 16S rRNA gene similarity (86.92%) with the Halanaerobiales, and represents at least a novel genus Anoxybacter within the class Clostridia. Here, we describe the first complete genome of strain DY22613T in this genus. The genome contains 3,557,532 bp with a G + C content of 35.88%. Genome sequence analysis of strain DY22613T reveals the presence of genes encoding riboflavin (ribBA,D,A,F,H), FAD­hydrogenases, etc. are involved in dissimilatory iron(III)-reducing process. The genome will provide insights into the mechanism of dissimilatory iron(III)-reducing in strain DY22613T, and contribute to better understand the role of this group in the biogeochemistry cycling of iron in deep-sea hydrothermal fields.

Extracellular electron transfer in fermentative bacterium Anoxybacter fermentans DY22613T isolated from deep-sea hydrothermal sulfides.

Dissimilatory Fe(III)-reducing bacteria (DIRBs) could reduce extracellular Fe(III) to Fe(II) via extracellular electron transfer (EET), playing an important role in biogeochemical cycling of Fe(III). Previous studies have noted the key role of multi-heme c-type cytochromes (MHCs) involved in EET by respiratory-type DIRBs, and proposed indirect electron transfer through the use of redox electron shuttles (e.g., flavins) or Fe(III)-chelation. However, knowledge about the EET of fermentative DIRBs was vitally scarce. Here, Anoxybacter fermentans DY22613T is a typical fermentative DIRB isolated from deep-sea hydrothermal sulfides, and it could utilize soluble Fe(III)-citrate and solid Fe(III)-bearing minerals as extracellular electron acceptors. Unlike respiratory-type DIRBs that utilize MHCs, this strain lacked MHCs to mediate EET. Besides, it did not adopt Fe(III)-chelation to mediate indirect EET. Nonetheless, genes encoding biosynthesis pathway of redox molecules (e.g., flavins) were found in its genome and their gene expression was up-regulated with Fe(III) reduction, suggesting redox molecules may mediate indirect EET by this strain. Subsequent physiological and biochemical tests further demonstrated endogenous riboflavin acted as main electron shuttles to mediate indirect EET by this strain, and menaquinone, indole played an assistant role in this process. Besides, this strain could employ exogenous humic acids to facilitate indirect EET. The mode of exogenous and endogenous redox molecules to co-mediate indirect EET by fermentative A. fermentans DY22613T, expands our knowledge about EET of fermentative DIRBs, and would contribute to better understand its ecological role in the biogeochemistry cycle of iron.

Importance of messenger RNA stability of toxin synthetase genes for monitoring toxic cyanobacterial bloom.

Toxic cyanobacterial blooms, occurring frequently worldwide, have posed serious threats to human health and aquatic ecosystem. RNA-based quantitative PCR, which could detect potential toxin-producing cyanobacteria that are actively transcribing toxin genes, is a more reliable method, compared to DNA-based qPCR. However, single-stranded mRNA is labile, and their degradation may lead to an underestimate of gene expression level, even misleading toxic risk management, and thus impeding its application. Here, the mRNA stability of microcystin synthetase genes (mcyA-J) was systematically evaluated in unicellular and colonial Microcystis with various treatments (-80 ℃, -196 ℃, 4 °C or 25 °C with RNases inhibitors). Results revealed the highly instability of toxin gene transcripts, affected by transcript structures and cell aggregation. The -196 ℃ treatment was the most effective for stabilizing these transcripts. RNAstore® (4 °C) could stabilize these transcripts effectively for a short time (less than 7 d), but their stability was strikingly reduced in colonial Microcystis. Furthermore, decay kinetics of mcyA-J transcripts in various treatments was developed, and showed that their decay rates were varied (0.0018-3.014 d-1), due to different molecular structures. The mcyH transcripts had the lowest decay rate (0.0018 d-1 at -196 ℃), attributed to the fewest AU sites and stem-loops involved in its secondary structure. Thus, mcyH was the most proper target gene for monitoring toxic cyanobacterial bloom. These findings provided new insight into mRNA stability of toxin genes, and contributed to monitoring toxic cyanobacterial blooms and water managements using RNA-based molecular techniques.

Evaluation of RNA degradation in pure culture and field Microcystis samples preserved with various treatments.

RNA-based molecular technique (RT-qPCR) is a promising method for microcystin monitoring in lakes and reservoirs, but great lability of RNA in cyanobacterial samples limits its application. To date, no studies have investigated how to effectively preserve RNA in cyanobacterial samples. In this study, four different treatments (-80 °C freezer, -196 °C liquid nitrogen, 4 °C or 25 °C preservation after adding RNA protective fluid) were employed to preserve RNA in pure culture and field Microcystis samples, and RNA degradation in these treatments were systematically evaluated. Results showed liquid nitrogen was the most effective treatment to preserve RNA in pure culture and field Microcystis samples. RNA preservation using RNA protective fluid was temperature dependent. Low temperature (4 °C) could effectively slow down RNA degradation within a short time (1-7 d), since decay rate of mcyH mRNA (k = 0.00094 d-1) was much lower at 4 °C than that at 25 °C (0.0549 d-1) (P < 0.05). However, for field samples, RNA degradation was much faster than pure culture samples with the same treatment. Therefore, to better preserve RNA in field samples, a practical strategy for RNA preservation combining RNA protective fluid and liquid nitrogen, was proposed. Tests of field experiments showed it was more effective than individual treatment for RNA preservation in Microcystis samples during field sampling. Thus, this strategy could be employed to preserve RNA in cyanobacterial samples during field sampling, which will contribute to the application of RT-qPCR technique for microcystin monitoring in lakes and reservoirs.