Ammonia nitrogen stress damages the intestinal mucosal barrier of yellow catfish (Pelteobagrus fulvidraco) and induces intestinal inflammation.

Nitrogen from ammonia is one of the most common pollutants toxics to aquatic species in aquatic environment. The intestinal mucosa is one of the key mucosal defenses of aquatic species, and the accumulation of ammonia nitrogen in water environment will cause irreversible damage to intestinal function. In this study, histology, immunohistochemistry, ultrastructural pathology, enzyme activity analysis and qRT-PCR were performed to reveal the toxic effect of ammonia nitrogen stress on the intestine of Pelteobagrus fulvidraco. According to histological findings, ammonia nitrogen stress caused structural damage to the intestine and reduced the number of mucous cells. Enzyme activity analysis revealed that the activity of bactericidal substances (Lysozyme, alkaline phosphatase, and ACP) had decreased. The ultrastructure revealed sparse and shortened microvilli as well as badly degraded tight junctions. Immunohistochemistry for ZO-1 demonstrated an impaired intestinal mucosal barrier. Furthermore, qRT-PCR revealed that tight junction related genes (ZO-1, Occludin, Claudin-1) were downregulated, while the pore-forming protein Claudin-2 was upregulated. Furthermore, as ammonia nitrogen concentration grew, so did the positive signal of Zap-70 (T/NK cell) and the expression of inflammation-related genes (TNF, IL-1ß, IL-8, IL-10). In light of the above findings, we conclude that ammonia nitrogen stress damages intestinal mucosal barrier of Pelteobagrus fulvidraco and induces intestinal inflammation.

The IL17 signaling pathway: A potential signaling pathway mediating gill hyperplasia and inflammation under ammonia nitrogen stress was identified by multi-omics analysis.

Ammonia nitrogen is extremely toxic to aquatic animals, and is also the most common pollutant in the aquatic environment. In order to investigate the effect of high concentration of ambient ammonia nitrogen on fish gills, two groups, including a high ammonia group (T group: TAN = 2.5 mg/L, 10 % 96 h LC50) and a control group (Z group: total ammonia nitrogen (TAN) = 0 mg/L) were set up in this study. The effects of chronic ammonia stress on the gills of Pelteobagrus fulvidraco were investigated by histopathological, enzymatic, transcriptomic and proteomic analyses after 28 d of stress at different ammonia nitrogen concentrations. Histopathological observations revealed significant inflammatory cell infiltration, necrotic and abscission at the base of the gill filaments, and massive proliferation of cells at the base of the gill lamellae. Ammonia nitrogen stress led to increased reactive oxygen species (ROS) content and decreased catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GSH-PX) activities in gills, indicating significant oxidative stress in gills. And further transcriptomic analysis revealed that 807 differential expression genes (DEGs) were generated in the gills, of which 587 DEGs were up-regulated and 220 DEGs were down-regulated. In addition, proteomics analysis identified 1073 differential expression proteins (DEPs) in gills, including 983 up- and 90 down-regulated DEPs. Pathway enrichment analysis of the DEGs and DEPs revealed that multiple inflammation-related signaling pathways were activated in the gill, including the significantly enriched IL17 signaling pathway. This suggests that IL17 signaling pathway might have a significant impact during signaling transduction. Further analysis of network regulation by mapping DEGs and DEPs to KEGG pathway revealed that IL17 signaling pathway mediated inflammation and cell proliferation in gills under ammonia stress. The results of this study provided new insights into the response of fish gills to ammonia nitrogen stress, and the IL17 signaling pathway may be a potential therapeutic target for reducing ammonia nitrogen gill toxicity.

Immunosuppression and apoptosis activation mediated by p53-Bcl2/Bax signaling pathway -The potential mechanism of goldfish (Carassius auratus Linnaeus) gill disease caused by Myxobolus ampullicapsulatus.

Myxobolus, a major harmful type of myxospora, is one of the main parasitic pathogens of freshwater fish. Once myxoboliosis occurs, treatment can be extremely difficult. Therefore, clear understandings of the etiology of myxoboliosis and its pathological mechanism are keys for prevention and control. Here, histology, transmission electron microscopy, transcriptome study, tunel assay, and immunohistochemistry were carried out, revealing the morphology, pathological effects as well as host response mechanism of goldfish gill to Myxobolus ampullicapsulatus. Histological studies showed that the mature spores of Myxobolus ampullicapsulatus were composed of three parts, the spore shell, sporoplasm and bottle shaped polar capsule containing double S-shaped polar filaments. Transcriptome analysis revealed that Myxobolus ampullicapsulatus -infected (Myx) goldfish gills were characterized by apoptosis activation mediated by "p53 signaling pathway" with significantly up-regulated apoptosis-related differential genes dominated by p53-Bcl2/Bax signaling pathway. In addition, tunel assay revealed severe gill apoptosis in the Myx group. Transcriptome analysis also revealed that Myx group showed changes in immune response and significantly down-regulated immune-related differential genes. Beyond that, immunohistochemistry showed that there was no significant increase in the number of gill lymphocyte after parasite infection. These results suggest that the pathological mechanism of Myxobolus ampullicapsulatus infection on gills of goldfish may be related to apoptosis and immunosuppression. Subsequent qRT-PCR showed that apoptosis-related genes (Caspase3,Bad, Bax) and anti-inflammatory gene IL-10 were significantly increased, while immune-related pro-inflammatory genes (IL-1ß, IL-8) were markedly down-regulated, further verifying the transcriptome results. Based on the above results, we concluded that p53-Bcl2/Bax related networks that dominant the expression of apoptosis genes were activated while immunity was suppressed in the gills of Myxobolus ampullicapsulatus infected goldfish. Our study is not only of benefit to enrich the taxonomy of Myxobolus but also clarifies its pathogenic mechanism, thus providing targets for prevention and control of myxoboliosis.

Measurement of methane emissions from CNG fueling stations in East China.

Methane, as the second most emitted greenhouse gas (GHG), has a warming potential of approximately 86 times that of carbon dioxide within 20 years. Quantifying methane emissions is helpful to the country's emission reduction efforts. However, currently, there is a lack of measurement data of methane emissions from natural gas fueling stations in China. In this study, a downwind quantification approach was employed to directly measure the methane emissions of nine compressed natural gas (CNG) fueling stations in East China according to the Environmental Protection Agency's Other Test Method 33A (OTM 33A). Moreover, methane concentrations were also measured near the nozzle of the refueling dispenser and the process equipment in the station. The methane emissions of the nine stations lied within the range of 0.11-0.83 kg/h, and the distribution of the emission rate was skewed. It was found that the emissions from gas fueling stations could be divided into intermittent emissions and continuous emissions, of which the intermittent emissions were the main source of methane.

cMOS enhanced the mucosal immune function of skin and gill of goldfish (Carassius auratus Linnaeus) to improve the resistance to Ichthyophthirius multifiliis infection.

METHODS: of supporting mucosal immune barrier integrity and prevention of some pathogenic infections in aquatic species, are key areas of active study, often focusing on feed additives. The objectives of this study were to explore the effects of feeding cMOS (concentrated mannan oligosaccharide) on the gill and skin mucosal barriers of goldfish (Carassius auratus Linnaeus) and evaluate health status during Ichthyophthirius multifiliis infection. After feeding the cMOS-containing diet for 60 days, Hematoxylin and eosin (H&E) staining showed greater length of gill lamella and thicker dermal dense layer, while Alcian Blue and Periodic acid-Schiff (AB-PAS) staining showed higher numbers of mucin cells in cMOS fed fish. Chemical analysis showed that fish fed cMOS had greater enzyme activity of lysozyme (LZM) and alkaline phosphatase (AKP) in gill and skin tissues, while qRT-PCR revealed higher expression of Muc-2 and IL-1ß, as well as lower expression of IL-10. After Ichthyophthirius multifiliis challenge, goldfish fed the cMOS diet had lower mortality and infection rates, as well as fewer visible white spots on the body surfaces. Histologically, the gill and skin of these fish presented less tissue damage and fewer parasites, and had a greater number of mucus cells. In addition, the expression of Muc-2 and IL-10 were notably higher while the expression of IL-1ß was significantly lower in cMOS fed goldfish than control fed fish. In this study, cMOS fed goldfish had stronger immune barrier function of skin and gill mucous, and better survival following Ichthyophthirius multifiliis infection.