Citizen science in action: Time-resolved immunofluorescence-based field detection of antibiotics with portable analytical kit.

Citizen scientist-based environmental monitoring and public education are becoming increasingly popular. However, current technologies for antibiotic-based novel contaminant identification are still restricted to laboratory sample collection and analysis due to detection methodologies and apparatus limitations. This study developed a time-resolved immunofluorescence-based simultaneous field-based assay for ciprofloxacin (CIP) and enrofloxacin (ENR) that matches test results to geographic locations. The assay helps the public understand the potential levels of antibiotic exposures in their environments and helps them take appropriate action to reduce risk. The assay was developed using smartphones and social software in addition to rapid testing. The method uses a portable, low-cost analytical kit with a smartphone app to build a field-based detection platform for the detection and analysis of ENR and CIP in water and aquatic products. The methodological evaluation was good, with detection limits of 0.4 ng/mL and 0.5 ng/g for ENR in water and fish, and quantification limits of 1.2 ng/mL and 1.4 ng/g, with recoveries of 89.0 %-101.0 % and 78.0 %-97.0 %. For CIP in water and fish, the limits of detection were 0.3 ng/mL and 0.4 ng/g, the limits of quantification were 0.9 ng/mL and 1.2 ng/g, and the recoveries were 75.0 %-91.0 % and 72.0 %-89.0 %, both with coefficients of variation <15 %. These limits were sufficient to prevent the two antibiotics from crossing over during simultaneous detection. The assay was validated using real samples to assess the effectiveness of the assay platform in field deployments, and the results were consistent with those obtained through liquid chromatography-tandem mass spectrometry (LC-MS) and enzyme-linked immunoassay (ELISA) techniques. In addition, the TRFIA assay process requires less time, uses more portable instruments, and is less complex than traditional methods. This study provides a new scientific, accurate, and rapid detection method for antibiotic detection by citizen scientists, helping scientists to obtain a wider range of data and providing more opportunities to solve scientific problems.

Assessment of the benefits of essential fatty acids and risks associated with antimicrobial residues in aquatic products: A case study of Chinese mitten crab (Eriocheir sinensis).

Much attention has been given to the safety and quality of aquatic products, including consuming Chinese mitten crab (Eriocheir sinensis), which offers both nutritional benefits and toxicological risks. Eighteen sulfonamides, 9 quinolones and 37 fatty acids were analyzed in 92 crab samples from primary aquaculture provinces in China. Enrofloxacin and ciprofloxacin have been mentioned as typical antimicrobials occurring at the greatest concentrations (>100 µg/kg, wm). By use of an in vitro method, the proportions of enrofloxacin, ciprofloxacin and essential fatty acids (EFAs, DHA and EPA) in ingested nutrients were determined to be 12 %, none and 95 %, respectively. The risk-benefit quotient (HQ) between the adverse effects of antimicrobials and nutritional benefits of EFAs in crabs found that HQs based on data after digestion were significantly less (HQ = 0.0086) than that of the control group where no digestion occurred (HQ = 0.055). This result suggested that (1) there was less risk posed by antimicrobials due to the consumption of crab, and (2) ignoring the bioaccessible fraction of antimicrobials in crabs might overestimate risks to the health of humans due to dietary exposure. Meaning bioaccessibility can improve the accuracy of the risk assessment process. Realistic risk evaluation should be recommended to achieve a quantified assessment of the dietary risks and benefits of aquatic products.

Quantitative detection of malachite green in sediment by a time-resolved immunofluorescence method combined with a portable 3D printing equipment platform.

Rapid detection technology of aquaculture fishery drug residues is needed to supplement large-scale instrument methods. To do this, the time-resolved fluorescence immunoassay (TRFIA) method and portable three-dimensional (3D) printing equipment platform were used, in combination with smartphones, to detect malachite green (MG) in pond sediments. The TRFIA was coupled to MG monoclonal antibodies (mAb) through lanthanide metal microspheres europium (Eu3+). The labeled antibody produced competitive immunity in the immune reaction system, and the specific fluorescence intensity in the product was determined by a portable 3D printing equipment platform to achieve quantitative analysis. To test this method, leucomalachite green (LMG) was converted to MG by oxidation of dicyanoquinone (DDQ), and a qualitative analysis was achieved. Methodological evaluation results were satisfactory, recoveries were 83 %-104 %, the limit of detection (LOD) was 0.3 ng/g, the limit of quantitation (LOQ) was 0.7 ng/g, and the coefficient of variation was 1.3 %-7.3 %. The linear equation y = -0.1496x + 0.5585 was in the range of 0-10 ng/g. The linear regression correlation coefficient was 99.2 %. The TRFIA was confirmed and positive samples were measured. Results were consistent with the standard method, which demonstrated that the TRFIA was feasible and that the detection results were reliable. Compared with the national standard method, the TRFIA saves time, is more convenient, and has high sensitivity. It provides an efficient technical method for the rapid screening of MG in the sediments of aquaculture environments.

Antibiotic accumulation, growth performance, intestinal diversification, and function of Nile tilapia (Oreochromis niloticus) feed by diets supplemented with different doses of sulfamethoxazole.

To comprehensively investigate the effects of exposure to legal doses of sulfamethoxazole (SMZ) in Nile tilapia (Oreochromis niloticus), fishes were exposed to diets supplemented with different doses of SMZ (NS, normal feed; LS, 20 mg/kg·day; MS, 200 mg/kg·day; and HS, 1000 mg/kg·day) for 4 weeks and then fed with normal feed for 4 weeks. General SMZ accumulation, growth performance, intestinal short-chain fatty acids, intestinal flora diversity, composition, and function were systemically evaluated. Results indicated that the SMZ accumulation in O. niloticus muscles, intestinal contents, and aquaculture environment positively correlated to the exposure dose. The growth performance, measured by weight increase, was MS>LS>NS, while HS antibiotics retarded the growth. SMZ-exposed O. niloticus had an increased number of fat particles in the liver and a change in the content of intestinal SCFAs. Moreover, SMZ exposure changed the biological diversity of the intestinal flora and subsequently induced microbiota dysbiosis, primarily inhibiting the growth of Fusobacteria, especially in HS group. Overall, exposure to higher SMZ doses than the recommended ones impair general intestinal functions and provokes health risk in fish. This study highlights the importance of rational and regulated use of SMZ in aquaculture.

Long-term exposure to environmental level of phenanthrene causes adaptive immune response and fibrosis in mouse kidneys.

As ubiquitous, persistent organic pollutants, polycyclic aromatic hydrocarbons (PAHs) have adverse impacts on human health. Phenanthrene (Phe) is one of the most abundant PAHs in the environment. However, the long-term effects of exposure to environmental level of Phe on the kidneys and the potential mechanisms are unclear. T helper (Th) cells, a subtype of CD4+ T cells that play a central role in the renal immune microenvironment. In this study, male mice were chronically exposed to 5, 50, and 500 ng/kg bw Phe every other day for total 210 days. Those results indicated that environmental Phe exposure caused kidney hypertrophy, injury and fibrosis in the mice. Chronic, long-term environmental level of Phe exposure did not significantly alter the innate immune response but induced adaptive immune response changes (Th1/Th2 related cytokines release), causing a type 1 immune response in the 5 ng/kg bw Phe group and a type 2 immune response in the high dose groups (50 and 500 ng/kg bw). This study provides novel insights into the roles of adaptive immune response in long-term PAH exposure-induced chronic kidney injury and fibrosis, which is beneficial for further understanding the potential health hazards of PAHs and providing new avenues for immune intervention strategies to alleviate PAHs toxicity.

Dynamic Elimination of Enrofloxacin Under Varying Temperature and pH in Aquaculture Water: An Orthogonal Study.

Orthogonal experiments were used to simulate the enrofloxacin (ENR) elimination dynamic in deeper water of aquaculture. Two factors at values common in fishery water (temperature of 20°C, 25°C, and 30°C; pH of 5, 7, and 9) were studied. The degradation of ENR in the nine treatment groups ranged from 44.7 to 80.1%. Variance analysis indicated that pH had a strong impact on the elimination of ENR, while temperature changes showed little effect. The ENR removal rate was highest at a combination of 25°C and pH 5. The optimal conditions of eliminating ENR were performed for exploring the generation of ciprofloxacin (CIP), which indicated that higher ENR concentrations led to the production of greater amounts of CIP. The half-time of ENR was increased 2.02-times in the ENR concentrations increasing from 20 to 2000 ng/mL. This study could increase our understanding of the behaviors of ENR and CIP during the aquaculture process.

Health risks associated with sulfonamide and quinolone residues in cultured Chinese mitten crab (Eriocheir sinensis) in China.

The occurrence of 27 antibiotics (18 sulfonamides and 9 quinolones) in 92 samples of cultured Chinese mitten crab (Eriocheir sinensis) from three provinces (Anhui, Jiangsu and Liaoning) was investigated. These 21 antibiotics were detected at least once in crabs from these provinces with detection frequencies of 3.70-90.91%. Sulfonamides were detected in 53.7% of the samples at concentrations of 0.1-10 µg/kg in Jiangsu, while quinolones were detected with 90.9% of samples containing concentrations of 1-100 µg/kg in Liaoning province. Enroflxacin, ciprofloxacin, sulfaquinoxaline, sulfameter, sulfadoxine, and sulfamethoxazole were the mainly used antibiotics and enroflxacin were present at a high concentration (>100 µg/kg). Dietary assessments showed that residual antibiotics in crabs from China were far below the maximum residue limit (MRL) of total sulfonamides and quinolones, and there was almost no risk associated with crab consumption. These results will provide meaningful indications for the safety of crab consumption.

Effects of fenvalerate concentrations and its chiral isomers on bacterial community structure in the sediment environment of aquaculture ponds.

To investigate the effect of chiral pesticide fenvalerate (FV) on the micro-ecological environment of aquaculture pond sediment, we used an indoor static experiment to observe the effects of FV added at different concentrations with different chiral isomers on the changes in the sediment bacterial community. The 16S rDNA high-throughput sequencing technique was used to conduct sequencing and analysis of the bacterial community structure as well as changes in aquaculture pond sediments after 4 weeks of cultivation. The results showed that the microbial alpha diversity indices (Sobs and Shannon indices) of the treated groups were significantly lower than those of the control group after 4 weeks (P < 0.05), and the values in the high-concentration group were significantly lower than those of the low-concentration group (P < 0.05). In terms of bacterial group composition, the proportion of abundance of Proteobacteria and Acidobacteria in the treated groups were greater than in the control group after 4 weeks, while the proportion of abundance of Bacteroidetes and Verrucomicrobia were lower. In the high-concentration FV treatment group, the proportion of abundance of Bacteroidetes, Acidobacteria, Chloroflexi, Nitrospinae, unclassified_k_norank, Ignavibacteriae, and Nitrospirae were significantly different from those of the other groups (P < 0.05). Principal coordinate analysis (PCoA) and ANONISIM/Adonis analysis showed that the cis-enantiomer had a stronger effect on the bacterial community as the concentration of FV increased. In addition, the linear discriminant analysis effect size (LEfSe) and linear discriminant analysis (LDA) results revealed differences in the level of enrichment of bacterial groups caused by FV at different concentrations and isomer levels. Collectively, this study showed that FV residue has a pronounced effect on bacterial communities in sediment, which becomes more significant with increasing exposure concentration. The effects of the cis- and trans-enantiomers of FV on the sediment environment are different; the cis-enantiomer has a stronger effect on the bacterial community.

Comparative study on nutrient removal of agricultural non-point source pollution for three filter media filling schemes in eco-soil reactors.

Nutrients (nitrogen and phosphorus) from agricultural non-point source (NPS) pollution have been increasingly recognized as a major contributor to the deterioration of water quality in recent years. The purpose of this article is to investigate the discrepancies in interception of nutrients in agricultural NPS pollution for eco-soil reactors using different filling schemes. Parallel eco-soil reactors of laboratory scale were created and filled with filter media, such as grit, zeolite, limestone, and gravel. Three filling schemes were adopted: increasing-sized filling (I-filling), decreasing-sized filling (D-filling), and blend-sized filling (B-filling). The systems were intermittent operations via simulated rainstorm runoff. The nutrient removal efficiency, biomass accumulation and vertical dissolved oxygen (DO) distribution were defined to assess the performance of eco-soil. The results showed that B-filling reactor presented an ideal DO for partial nitrification-denitrification across the eco-soil, and B-filling was the most stable in the change of bio-film accumulation trends with depth in the three fillings. Simultaneous and highest removals of NH4(+)-N (57.74-70.52%), total nitrogen (43.69-54.50%), and total phosphorus (42.50-55.00%) were obtained in the B-filling, demonstrating the efficiency of the blend filling schemes of eco-soil for oxygen transfer and biomass accumulation to cope with agricultural NPS pollution.

Propagule Pressure, Habitat Conditions and Clonal Integration Influence the Establishment and Growth of an Invasive Clonal Plant, Alternanthera philoxeroides.

Many notorious invasive plants are clonal, spreading mainly by vegetative propagules. Propagule pressure (the number of propagules) may affect the establishment, growth, and thus invasion success of these clonal plants, and such effects may also depend on habitat conditions. To understand how propagule pressure, habitat conditions and clonal integration affect the establishment and growth of the invasive clonal plants, an 8-week greenhouse with an invasive clonal plant, Alternanthera philoxeroides was conducted. High (five fragments) or low (one fragment) propagule pressure was established either in bare soil (open habitat) or dense native vegetation of Jussiaea repens (vegetative habitat), with the stolon connections either severed from or connected to the relatively older ramets. High propagule pressure greatly increased the establishment and growth of A. philoxeroides, especially when it grew in vegetative habitats. Surprisingly, high propagule pressure significantly reduced the growth of individual plants of A. philoxeroides in open habitats, whereas it did not affect the individual growth in vegetative habitats. A shift in the intraspecific interaction on A. philoxeroides from competition in open habitats to facilitation in vegetative habitats may be the main reason. Moreover, clonal integration significantly improved the growth of A. philoxeroides only in open habitats, especially with low propagule pressure, whereas it had no effects on the growth and competitive ability of A. philoxeroides in vegetative habitats, suggesting that clonal integration may be of most important for A. philoxeroides to explore new open space and spread. These findings suggest that propagule pressure may be crucial for the invasion success of A. philoxeroides, and such an effect also depends on habitat conditions.