Physiological and metabolic toxicity of polystyrene microplastics to Dunaliella salina.

The toxicity of microplastics (MPs) to marine microalgae has raised much concern. However, research at metabolic level is quite limited. In this study, the physiological and metabolic effects of polystyrene (PS) and aged polystyrene (A-PS) MPs on Dunaliella salina were investigated. Both PS and A-PS inhibited the growth of microalgae, but promoted the pigment synthesis in algal cells. The oxidative stress analysis indicated that PS and A-PS induced high production of reactive oxygen species (ROS), and caused oxidative damage to algal cells. Particularly, the highest ROS level in PS and A-PS groups were 1.70- and 2.24-fold of that in the control group, respectively. Untargeted metabolomics analysis indicated that PS and A-PS significantly increased the differential metabolites. Compared with the control group, the significant upregulation of glycerophospholipids metabolites illustrated that severe membrane lipid peroxidation occurred in the MPs groups. Metabolic pathways analysis showed that PS and A-PS perturbed the amino acid-related metabolic pathways. In particular, the amino acid biosynthesis and ATP-binding cassette (ABC) transporter pathways were significantly upregulated, thus promoting nitrogen storage and transmembrane transport in Dunaliella salina. Transmembrane transport requires a large amount of ATP; as a result, algal cell division is inhibited. In addition, A-PS stimulated more active glutathione metabolism than PS. These results enrich the understanding of the toxicity of PS MPs to microalgae at the metabolic level, and are helpful for further assessing the ecological impacts of MPs on microalgae.

Single and combined toxicity of polystyrene nanoplastics and copper on Platymonas helgolandica var. tsingtaoensis: Perspectives from growth inhibition, chlorophyll content and oxidative stress.

The combined toxic effects of nanoplastics and heavy metals on aquatic organisms have attracted widespread attention; however, the results are inconsistent and the mechanisms remain unclear. In this study, the single and combined toxicity effects of Cu and two types of polystyrene nanoplastics (PS-NPs; 50 nm PS and 55 nm PS-COOH) on Platymonas helgolandica var. tsingtaoensis were investigated, including growth inhibition, chlorophyll content, and oxidative stress. An adverse dose-response relationship on growth inhibition was found in the Cu treatment groups, which was related to the decrease in chlorophyll content and damage to cell membranes. The growth inhibitory effect of PS-NPs on microalgae increased with exposure time and concentration, and no significant difference was found in the two types of PS-NPs because of the negligible contribution of functional groups. A more significant increase in chlorophyll content was found in PS treatments than in PS-COOH treatments at 96 h because of the microscale aggregates formed by PS. Higher concentrations (≥ 50 mg/L) of PS-NPs caused membrane lipid peroxidation, which might be responsible for growth inhibition. In the combined exposure experiments, a synergistic effect on the growth inhibition rate was obtained using the independent action model and Abbott model. Combined exposure triggered more severe oxidative damage to the microalgae. Adsorption experiment results showed that there was no adsorption between PS-NPs and Cu, while the interaction of Cu and algal cells could be promoted due to the presence of the PS-NPs, which explained the increasing combined toxicity. This study could improve our understanding of the combined toxicity of nanoplastics and heavy metals and could provide a new explanation for the mechanism of combined toxicity.

Effects of Nanoplastics on the Dinoflagellate Amphidinium carterae Hulburt from the Perspectives of Algal Growth, Oxidative Stress and Hemolysin Production.

Recently, the effects of nanoplastics (NPs) on aquatic organisms have attracted much attention; however, research on the toxicity of NPs to microalgae has been insufficient. In the present study, the effects of polystyrene nanoplastics (nano-PS, 50 nm) on growth inhibition, chlorophyll content, oxidative stress, and algal toxin production of the marine toxigenic dinoflagellate Amphidinium carterae Hulburt were investigated. Chlorophyll synthesis was promoted by nano-PS on day 2 but was inhibited on day 4; high concentrations of nano-PS (≥50 mg/L) significantly inhibited the growth of A. carterae. Moreover, despite the combined effect of superoxide dismutase (SOD) and glutathione (GSH), high reactive oxygen species (ROS) level and malondialdehyde (MDA) content were still induced by nano-PS (≥50 mg/L), indicating severe lipid peroxidation. In addition, the contents of extracellular and intracellular hemolytic toxins in nano-PS groups were significantly higher than those in control groups on days 2 and 8, except that those of extracellular hemolytic toxins in the 100 mg/L nano-PS group decreased on day 8 because of severe adsorption of hemolytic toxins to the nano-PS. Hence, the effects of nano-PS on A. carterae are closely linked to nano-PS concentration and surface properties and exposure time. These findings provide a deep understanding of the complex effects of NPs on toxigenic microalgae and present valuable data for assessing their environmental risks.

Phototransformation of Graphene Oxide on the Removal of Sulfamethazine in a Water Environment.

Graphene oxide (GO) is widely used in various fields and has raised concerns regarding its potential environmental fate and effect. However, there are few studies on its influence on coexisting pollutants. In this study, the phototransformation of GO and coexisting sulfamethazine (SMZ) under UV irradiation was investigated, with a focus on the role of reactive oxygen species. The results demonstrated that GO promoted the degradation of SMZ under UV irradiation. The higher the concentration of GO, the higher the degradation rate of SMZ, and the faster the first-order reaction rate. Two main radicals, ∙OH and 1O2, both contributed greatly in terms of regulating the removal of SMZ. Cl-, SO42-, and pH mainly promoted SMZ degradation by increasing the generation of ∙OH, while humic acid inhibited SMZ degradation due to the reduction of ∙OH. Moreover, after UV illumination, the GO suspension changed from light yellow to dark brown with increasing absorbance at a wavelength of 225 nm. Raman spectra revealed that the ID/IG ratio slightly decreased, indicating that some of the functional groups on the surface of GO were removed under low-intensity UV illumination. This study revealed that GO plays important roles in the photochemical transformation of environmental pollutants, which is helpful for understanding the environmental behaviors and risks of nanoparticles in aquatic environments.

Effects of polystyrene and triphenyl phosphate on growth, photosynthesis and oxidative stress of Chaetoceros meülleri.

The toxicity of microplastics to marine organisms has attracted much attention; however, studies of their effects on marine microalgae remain limited. Here, the effects of the single and combined toxicity of polystyrene (PS) and triphenyl phosphate (TPhP) on the cell growth, photosynthesis, and oxidative stress of Chaetoceros meülleri were investigated. PS inhibited growth of the algae cells and caused a dose-dependent effect on oxidative stress. The significantly high production of reactive oxygen species (ROS) induced severe cell membrane damage, as confirmed by high fluorescence polarization. However, there was no obvious decrease in chlorophyll a content, and 80 mg/L of PS significantly promoted chlorophyll a synthesis. The TPhP also inhibited cell growth, except at low concentrations (0.2-0.8 mg/L), which stimulated algae growth over 48 h. Moreover, no obvious decrease in chlorophyll a and maximal photochemical efficiency of PSII was found in the TPhP experimental groups except for 3.2 mg/L TPhP, where the rapid light curves showed a significantly reduced photosynthetic capacity of algae. In addition, TPhP caused high ROS levels at 96 h, resulting in cell membrane damage. Using the additive index and independent action methods, the combined toxic effects of PS and TPhP on the algae were evaluated as antagonistic; however, cell membrane damage caused by high ROS levels was still noticeable. This study has shown the potential toxicity of PS and TPhP to marine microalgae, and provided insights into the combined risk assessment of TPhP and microplastics in the marine environment.

A set of RT-PCR assays for detection of all known avian paramyxoviruses and application in surveillance of avian paramyxoviruses in China.

BACKGROUND: Avian paramyxoviruses (APMVs), also termed avian avulaviruses, are of a vast diversity and great significance in poultry. Detection of all known APMVs is challenging, and distribution of APMVs have not been well investigated. METHODS: A set of reverse transcription polymerase chain reaction (RT-PCR) assays for detection of all known APMVs were established using degenerate primers targeting the viral polymerase L gene. The assays were preliminarily evaluated using in-vitro transcribed double-stranded RNA controls and 24 known viruses, and then they were employed to detect 4,346 avian samples collected from 11 provinces. RESULTS: The assays could detect 20-200 copies of the double-stranded RNA controls, and detected correctly the 24 known viruses. Of the 4,346 avian samples detected using the assays, 72 samples were found positive. Of the 72 positives, 70 were confirmed through sequencing, indicating the assays were specific for APMVs. The 4,346 samples were also detected using a reported RT-PCR assay, and the results showed this RT-PCR assay was less sensitive than the assays reported here. Of the 70 confirmed positives, 40 were class I Newcastle disease virus (NDV or APMV-1) and 27 were class II NDV from poultry including chickens, ducks, geese, and pigeons, and three were APMV-2 from parrots. The surveillance identified APMV-2 in parrots for the first time, and revealed that prevalence of NDVs in live poultry markets was higher than that in poultry farms. The surveillance also suggested that class I NDVs in chickens could be as prevalent as in ducks, and class II NDVs in ducks could be more prevalent than in chickens, and class II NDVs could be more prevalent than class I NDVs in ducks. Altogether, we developed a set of specific and sensitive RT-PCR assays for detection of all known APMVs, and conducted a large-scale surveillance using the assays which shed novel insights into APMV epidemiology.

Phototransformation of zinc oxide nanoparticles and coexisting pollutant: Role of reactive oxygen species.

In this study, the photochemistry of ZnO NPs and their effect on phototransformation of coexisting pollutants (sulfamethazine, SMZ) were systematically investigated under UV illumination. SMZ (40 µM) degradation was accelerated by ZnO NPs, as the observed reaction rate constant (kobs) increased from 0.0809 h-1 to 0.7982 h-1 at the concentration of 5-50 mg/L ZnO NPs. Free radical quenching and quantification experiments indicated the reactive oxygen species, especially the hydroxyl radicals (OH) and singlet oxygen (1O2), made great contributions to SMZ degradation. Moreover, SMZ was prone to be degraded at high pH with kobs reaching upto 0.5734 h-1 at pH 12.0. The presence of Cl- (1000 mM) reduced the SMZ decomposition greatly by 2.4-fold while the effects of SO42- (30 mM) were very limited. Natural organic matter including humic acid and tannic acid both inhibited the degradation of SMZ with kobs decreasing by 35.4-fold and 132-fold, respectively. During the photoreaction process, ZnO NPs fragmented into relative small size pieces obviously along with the release of Zn2+. Finally, the possible cotransformation pathways of ZnO NPs and SMZ were proposed based on SMZ degradation intermediates and the above results. These findings of the present study suggested that the photoreactions of ZnO NPs greatly influenced the transformation of contaminants and ZnO NPs themselves in aquatic environment, which may have significant implications for the fate assessment of NPs and environmental pollutants.

Sorption of fluoroquinolones to nanoplastics as affected by surface functionalization and solution chemistry.

Microplastics have attracted much attention in recent years as they can interact with pollutants in water environment. However, nanoplastics (NPs) with or without the surface functionalization modification have not been thoroughly explored. Here, the sorption behaviors of two fluoroquinolones (FQs), including norfloxacin (NOR) and levofloxacin (LEV) on polystyrene NPs (nano-PS) and carboxyl-functionalized polystyrene NPs (nano-PS-COOH) were investigated. The results showed that sorption isotherms were nonlinear and well fitted by Langmuir model. The sorption capacities of NOR and LEV on nano-PS-COOH were higher than those on nano-PS, and their physical interactions, including polar interaction, electrostatic interaction and hydrogen bonding may be the dominant mechanisms. Moreover, the increase of pH firstly increased the sorption of two FQs on NPs and then decreased because NOR and LEV had a reverse charge at different pH values. Salinity and dissolved organic matter both inhibited the sorption process. These findings show that NPs with or without the surface functionalization modification have different sorption behaviors for environmental pollutants, which deserve our further concern.

Effects of oxidation degree on photo-transformation and the resulting toxicity of graphene oxide in aqueous environment.

Graphene oxide (GO) has been demonstrated to be key component for diverse applications. However, their potential environmental reactivity, fate and risk have not been fully evaluated to date. In this study, we investigated the photochemical reactivity of four types of GO with different oxidation degrees in aqueous environment, and their related toxicity to two bacterial models Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) was further compared. After UV-irradiation, a large amount of oxygen functional groups on GO were reduced and the electronic conjugations within GO were restored as indicated by UV-visible absorption spectra, X-ray photoelectron spectroscopy and Raman spectroscopy analysis. Moreover, the higher the oxidation degree of the pristine GO was, the more obvious of the photo-transformation changes were. In order to further reveal the photochemical reactivity mechanisms, the reactive oxygen species (ROS) generation of GO was monitored. The quantity of ROS including singlet oxygen (1O2), superoxide anions (O2·-), and hydroxyl radicals (·OH) increased with increasing oxidation degree of GO, which was in accordance with the previous characterization results. Scanning electron microscopy and cell growth analyses of E. coli and S. aureus showed that the photochemical transformation enhanced the toxicity of GO, which might be due to an increase in functional group density. The higher conductivity of the reduced graphene oxide (RGO) was responsible for its stronger toxicity than GO through membrane damage and oxidative stress to bacteria. This study revealed that the oxidation degrees play important roles in photochemical transformation and the resulting toxicity of GO, which is helpful for understanding the environmental behaviors and risks of GO in aquatic environments.

Genome sequence characterization of canine parvoviruses prevalent in the Sichuan province of China.

Canine parvovirus 2 (CPV-2) infection is responsible for large numbers of animal deaths worldwide and is one of the most dangerous infectious diseases in young puppies. Twenty-four rectal swabs were collected from dogs with clinical signs of vomiting and haemorrhagic diarrhoea and were initially verified to be infected with CPV-2 using colloidal gold test strips. From the 24 CPV-positive samples, complete genome of 5050-5054 nucleotides was sequenced with a next-generation sequencing platform. Characteristics of the Open Reading Frames from different CPV-2 strains detected in this study were analyzed. Several VP2 point mutations were discovered, and demonstrated the co-circulation of new CPV-2a, new CPV-2b and CPV-2c in Sichuan province of China. The analysis results of the Chinese CPV-2 retrieved from the NCBI nucleotide, showed that new CPV-2a has become the predominant variant in some provinces of China. Phylogenetic analysis of global VP2 and NS1 nucleotide sequences revealed certain correlations among geographical regions, types and circulating time, which lays the foundation for further research concerning the epidemiology, genetic variation, vaccination and molecular evolutionary relationships of the CPV-2 identified at different times and from different regions.

Interactions between microplastics and phthalate esters as affected by microplastics characteristics and solution chemistry.

Microplastics have become a major concern in recent years as they can be recognized as the transport vectors for pollutants in environment. In this study, the sorption behavior of two phthalate esters (PAEs), including diethyl phthalate (DEP) and dibutyl phthalate (DBP), onto three types of microplastics (PVC: polyvinyl chloride, PE: polyethylene, and PS: polystyrene) was investigated. The sorption isotherms of both DEP and DBP on microplastics were highly linear, suggesting that the partition was the main sorption mechanism. The Kd values of DBP were much higher than those of DEP, demonstrating that hydrophobic interaction governed the partition mechanism. Sorption of the two PAEs on the three microplastics followed the order of PS > PE > PVC, indicating that chemical properties of microplastics played an important roles in their sorption behaviors. Solution pH and natural organic matter had no significant impact on PAEs sorption by microplastics. However, the presence of NaCl and CaCl2 enhanced the sorption of both DEP and DBP because of the salting-out effect. The findings of the present study may have significant implications for the fate and transport assessment of both PAEs and microplastics.

Joint toxicity of microplastics with triclosan to marine microalgae Skeletonema costatum.

Toxicity of single microplastics on organisms has been reported widely, however, their joint toxicity with other contaminants on phytoplankton is rarely investigated. Here, we studied the toxicity of triclosan (TCS) with four kinds of microplastics namely polyethylene (PE, 74 µm), polystyrene (PS, 74 µm), polyvinyl chloride (PVC, 74 µm), and PVC800 (1 µm) on microalgae Skeletonema costatum. Both growth inhibition and oxidative stress including superoxide dismutase (SOD) and malondialdehyde (MDA) were determined. We found that TCS had obvious inhibition effect on microalgae growth within the test concentrations, and single microplastics also had significant inhibition effect which followed the order of PVC800 > PVC > PS > PE. However, the joint toxicity of PVC and PVC800 in combination with TCS decreased more than that of PE and PS. The higher adsorption capacity of TCS on PVC and PVC800 was one possible reason for the greater reduction of their toxicity. The joint toxicity of PVC800 was still most significant (PE < PVC < PS < PVC800) because of the minimum particle size. According to the independent action model, the joint toxicity systems were all antagonism. Moreover, the reduction of SOD was higher than MDA which revealed that the physical damage was more serious than intracellular damage. SEM images revealed that the aggregation of microplastics and physical damage on algae was obvious. Collectively, the present research provides evidences that the existence of organic pollutants is capable of influencing the effects of microplastics, and the further research on the joint toxicity of microplastics with different pollutants is urgent.

Comparison of three media for transport and storage of the samples collected for detection of avian influenza virus.

Detection of avian influenza viruses (AIVs) is important for diagnosis, surveillance and control of avian influenza which is of great economic and public health significance. Proper transport and storage of samples is critical for the detection when the samples cannot be detected immediately. As recommended by some international or national authoritative entities and some publications, phosphate buffered saline (PBS), PBS-glycerol and brain heart infusion broth (BHIB) are frequently used for transport and storage of the samples collected for detection of AIVs worldwide. In this study, we compared these three media for transport and storage of simulated and authentic swab and feces samples collected for detection of AIVs using virus isolation and reverse transcription-PCR. The results suggest that PBS-glycerol is superior to PBS and BHIB as the sample transport and storage media. The results also suggest that the samples collected for detection of AIVs should be detected as soon as possible because the virus concentration of the samples may decline rapidly during storage within days at 4 or -20°C.

Genomic Analysis and Surveillance of the Coronavirus Dominant in Ducks in China.

The genetic diversity, evolution, distribution, and taxonomy of some coronaviruses dominant in birds other than chickens remain enigmatic. In this study we sequenced the genome of a newly identified coronavirus dominant in ducks (DdCoV), and performed a large-scale surveillance of coronaviruses in chickens and ducks using a conserved RT-PCR assay. The viral genome harbors a tandem repeat which is rare in vertebrate RNA viruses. The repeat is homologous to some proteins of various cellular organisms, but its origin remains unknown. Many substitutions, insertions, deletions, and some frameshifts and recombination events have occurred in the genome of the DdCoV, as compared with the coronavirus dominant in chickens (CdCoV). The distances between DdCoV and CdCoV are large enough to separate them into different species within the genus Gammacoronavirus. Our surveillance demonstrated that DdCoVs and CdCoVs belong to different lineages and occupy different ecological niches, further supporting that they should be classified into different species. Our surveillance also demonstrated that DdCoVs and CdCoVs are prevalent in live poultry markets in some regions of China. In conclusion, this study shed novel insight into the genetic diversity, evolution, distribution, and taxonomy of the coronaviruses circulating in chickens and ducks.

Subclinical highly pathogenic avian influenza virus infection among vaccinated chickens, China.

Subclinical infection of vaccinated chickens with a highly pathogenic avian influenza A(H5N2) virus was identified through routine surveillance in China. Investigation suggested that the virus has evolved into multiple genotypes. To better control transmission of the virus, we recommend a strengthened program of education, biosecurity, rapid diagnostics, surveillance, and elimination of infected poultry.

Evaluation of avian influenza virus isolated from ducks as a potential live vaccine candidate against novel H7N9 viruses.

Recent outbreaks of a novel H7N9 avian influenza virus in humans in China raise pandemic concerns and underscore an urgent need to develop effective vaccines. Theoretically, live influenza vaccines are of multiple advantages over traditional inactivated influenza vaccines to be used in a pandemic, because they can be produced rapidly, safely, and inexpensively. However, studies on live vaccines against the novel H7N9 virus are limited. In this study, we evaluated a potential live influenza vaccine candidate using an H7N3 avian influenza virus isolated from ducks with controls of two recombinant viruses generated through reverse genetics. The potential candidate could be produced efficiently using chicken embryonated eggs, and is homogenous to the novel H7N9 virus in their viral hemagglutinin genes. The potential candidate is likely low pathogenic to birds and mammals, and likely sensitive to oseltamivir and amantadine, as suggested by its genomic sequences. Its low pathogenicity was further supported through inoculation in mice, chicken embryonated eggs and chickens. Specific antibodies elicited in mice were detectable at least during the period between day 14 and day 56 after intranasal administration of the candidate for one time. Titers of the specific antibodies increased significantly with a boost intranasal administration or a higher inoculation dose. The induced specific antibodies were of substantial cross-reactivity with the novel H7N9 virus. These primary but promising evaluation data suggest that the duck influenza virus could be used as a potential live vaccine candidate, favorably through a prime-boost route, to mitigate the severity of the possible pandemic caused by the newly emerging H7N9 virus, and is valuable to be further evaluated.

Identification of a potential novel type of influenza virus in Bovine in China.

Bovine influenza virus was first identified in the USA in 2013, and the virus represents a potential novel type of influenza viruses. However, the distribution and evolution of the virus remain unknown. We conducted a pilot survey of bovine influenza virus in China, and identified three bovine influenza viruses which are highly homogenous to the ones identified in the USA, suggesting that the bovine influenza virus likely circulates widely and evolves slowly in the world.

A clinical survey of common avian infectious diseases in China.

Multiple common avian infectious diseases (CAIDs), namely, avian infectious diseases excluding highly pathogenic avian influenza and Newcastle disease, such as avian salmonellosis and coccidiosis, cause huge economic loss in poultry production and are of great significance in public health. However, they are usually not covered in the systems for reporting of animal diseases. Consequently, the distribution of CAIDs is not clear in many countries. Here, we report a clinical survey of CAIDs in China based on clinical diagnosis of eight veterinary clinics in 2011 and 2012. This survey provided the distribution data of viral, bacterial, and parasitic CAIDs in different types of avian flocks, seasons, and regions, data that are of great value in the research, prevention, and control of poultry diseases. This survey suggested that avian colibacillosis, infectious serositis in ducks caused by Riemerella anatipestifer, avian salmonellosis, fowl cholera, avian mycoplasmosis, avian aspergillosis, coccidiosis, low pathogenic avian influenza, infectious bronchitis, infectious bursal disease, and infectious laryngotracheitis are likely to be prevalent in the poultry in China.

[Expression and significance of tumor susceptibility gene 101 in hepatocellular carcinoma tissues].

AIM: To study the expression and clinical significance of tumor susceptibility gene 101 (TSG101) expression in hepatocellular carcinoma (HCC) tissues. METHODS: The expression of TSG101 in HCC and corresponding non-tumor tissues was detected by immunohistochemistry and Western blotting. Statistical analyses were conducted to test the relationships of TSG101 expression with clinical parameters such as age, gender, TNM stage, tumor metastasis, and so on. RESULTS: Immunohistochemistry and Western blotting showed that the expression of TSG101 in HCC was significantly higher than that in corresponding non-tumor tissues (P<0.05), and the expression rate was also higher in HCC (53/66, 80.3%) than in non-cancer tissues(18/66, 20.7%)(P<0.05). Higher TSG101 expression in HCC was significantly correlated with TNM stage (P<0.05) and metastasis (P<0.05), but not with age, gender and HBsAg (P>0.05). Multiple regression analysis indicated that TSG101 expression rate was significantly associated with TNM stage and metastasis (P<0.05), but not with gender, age and HBsAg (P>0.05). CONCLUSION: The expression of TSG101 in HCC is higher than that in corresponding non-cancer tissues and the expression level is closely correlated with TNM stage and metastasis of HCC.