Ultrathin Self-Healing Nanofibrous Membrane with a Hierarchical Confined Structure for Biomimetic Epidermal Electrodes.

Integrating self-healing capabilities into epidermal electrodes is crucial to improving their reliability and longevity. Self-healing nanofibrous materials are considered an ideal candidate for constructing ultrathin, long-lasting wearable epidermal electrodes due to their lightweight and high breathability. However, due to the strong interaction between fibers, self-healing nanofiber membranes cannot exist stably. Therefore, the development of self-healing and breathable nanofibrous epidermal electrodes still remains a major challenge. Here, a hierarchical confinement strategy that combines molecular and spatial confinement to overcome supramolecular hydrogen bonding between self-healing nanofibers is reported, and an ultrathin self-healing nanofibrous epidermal electrode with a neural net-like structure is developed. It can achieve real-time monitoring of electrophysiological signals through long-term conformal attachment to skin or plants and has no adverse effects on skin health or plant growth. Given the almost imperceptible nature of epidermal electrodes to users and plants, it lays the foundation for the development of biocompatible, self-healing, wearable, flexible electronics.

Impairment of oocyte quality caused by gut microbiota dysbiosis in obesity.

Obesity poses risks to oocyte maturation and embryonic development in mice and humans, linked to gut microbiota dysbiosis and altered host metabolomes. However, it is unclear whether symbiotic gut microbes have a pivotal role in oocyte quality. In mouse models of fecal microbiota transplantation, we demonstrated aberrant meiotic apparatus and impaired maternal mRNA in oocytes, which is coincident with the poor developmental competence of embryos. Using metabolomics profiling, we discovered that the cytosine and cytidine metabolism was disturbed, which could account for the fertility defects observed in the high-fat diet (HFD) recipient mice. Additionally, cytosine and cytidine are closely related with gut microbiota dysbiosis, which is accompanied by a notable reduction of abundance of Christensenellaceae R-7 group in the HFD mice. In summary, our findings provided evidence that modifying the gut microbiota may be of value in the treatment of infertile female individuals with obesity.

Intracranial hypertension as the primary symptom of malignant melanoma: A case report.

BACKGROUND: The etiological diagnosis of intracranial hypertension is quite complicated but important in clinical practice. Some common causes are craniocerebral injury, intracranial space-occupying lesion, subarachnoid hemorrhage, and hydrocephalus. When a patient presents with intracranial hypertension, the common causes are to be considered first so that other causes would be dismissed. With the morbidity lower than 9%, neuromelanin is very rare. Common symptoms include nerve damage symptoms, epilepsy, psychiatric symptoms, and cognitive disorders. CASE SUMMARY: We present a patient with melanoma which manifested with isolated intracranial hypertension without any other neurological signs. A 22-year-old male had repeated nausea and vomiting for 2 mo with Babinski sign (+) on both sides, nuchal rigidity, and subarachnoid hemorrhage. He had been diagnosed with melanoma and was given surgery and whole-brain radiation. Ultimately, the patient died 2 mo later. CONCLUSION: Malignant melanoma should be taken into consideration in the differential diagnosis of intracranial hypertension.

SMARCA2 and SMARCA4 Participate in DNA Damage Repair.

BACKGROUND: The switching/sucrose non-fermentable (SWI/SNF) Related, Matrix Associated, Actin Dependent Regulator Of Chromatin, Subfamily A (SMARCA) member 2 and member 4 (SMARCA2/4) are paralogs and act as the key enzymatic subunits in the SWI/SNF complex for chromatin remodeling. However, the role of SMARCA2/4 in DNA damage response remains unclear. METHODS: Laser microirradiation assays were performed to examine the key domains of SMARCA2/4 for the relocation of the SWI/SNF complex to DNA lesions. To examine the key factors that mediate the recruitment of SMARCA2/4, the relocation of SMARCA2/4 to DNA lesions was examined in HeLa cells treated with inhibitors of Ataxia-telangiectasia-mutated (ATM), Ataxia telangiectasia and Rad3-related protein (ATR), CREB-binding protein (CBP) and its homologue p300 (p300/CBP), or Poly (ADP-ribose) polymerase (PARP) 1/2 as well as in H2AX-deficient HeLa cells. Moreover, by concomitantly suppressing SMARCA2/4 with the small molecule inhibitor FHD286 or Compound 14, the function of SMARCA2/4 in Radiation sensitive 51 (RAD51) foci formation and homologous recombination repair was examined. Finally, using a colony formation assay, the synergistic effect of PARP inhibitors and SMARCA2/4 inhibitors on the suppression of tumor cell growth was examined. RESULTS: We show that SMARCA2/4 relocate to DNA lesions in response to DNA damage, which requires their ATPase activities. Moreover, these ATPase activities are also required for the relocation of other subunits in the SWI/SNF complex to DNA lesions. Interestingly, the relocation of SMARCA2/4 is independent of γH2AX, ATM, ATR, p300/CBP, or PARP1/2, indicating that it may directly recognize DNA lesions as a DNA damage sensor. Lacking SMARCA2/4 prolongs the retention of γH2AX, Ring Finger Protein 8 (RNF8) and Breast cancer susceptibility gene 1 (BRCA1) at DNA lesions and impairs RAD51-dependent homologous recombination repair. Furthermore, the treatment of an SMARCA2/4 inhibitor sensitizes tumor cells to PARP inhibitor treatment. CONCLUSIONS: This study reveals SMARCA2/4 as a DNA damage repair factor for double-strand break repair.

Immune features revealed by single-cell RNA and single-cell TCR/BCR sequencing in patients with rheumatoid arthritis receiving COVID-19 booster vaccination.

Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, have profoundly affected human health. Booster COVID-19 vaccines have demonstrated significant efficacy in reducing infection and severe cases. However, the effects of booster COVID-19 vaccines on key immune cell subsets and their responses in rheumatoid arthritis (RA) are not well understood. By using single-cell RNA sequencing (scRNA-seq) combined with scTCR/BCR-seq analysis, a total of 8 major and 27 minor cell clusters were identified from paired peripheral blood mononuclear cells (PBMCs) which were collected 1 week before and 4 weeks after booster vaccination in stable RA patients. Booster vaccination only had limited impact on the composition and proportions of PBMCs cell clusters. CD8+ cytotoxic T cells (CD8+T_CTL) showed a trend toward an increase after vaccination, while naive B cells and conventional dendritic cells (cDCs) showed a trend toward a decrease. Transcriptomic changes were observed after booster vaccination, primarily involving T/B cell receptor signaling pathways, phagosome, antigen processing and presenting, and viral myocarditis pathways. Interferon (IFN) and pro-inflammatory response gene sets were slightly upregulated across most major cell subpopulations in COVID-19 booster-vaccinated RA individuals. Plasma neutralizing antibody titers significantly increased after booster COVID-19 vaccination (p = 0.037). Single-cell TCR/BCR analysis revealed increased B cell clone expansion and repertoire diversity postvaccination, with no consistent alterations in T cells. Several clonotypes of BCRs and TCRs were identified to be significantly over-represented after vaccination, such as IGHV3-15 and TRBV28. Our study provided a comprehensive single-cell atlas of the peripheral immune response and TCR/BCR immune repertoire profiles to inactivated SARS-CoV-2 booster vaccination in RA patients, which helps us to understand vaccine-induced immune responses better.

Aperture division multispectral camera for the Earth's reflected solar radiation observation based on the Lagrange L1 point of the Earth-Moon system.

We propose an aperture division multispectral camera for Earth observation (EAMC) based on the Lagrange L1 point of the Earth-Moon system to measure the Earth's reflected solar radiation (RSR), quantify the effective radiative forcing (ERF) and establish the pixel-scale multispectral angular distribution model (ADM) of the Earth's radiance. The EAMC adopts the snapshot technique to provide multispectral images in the 360-920 nm wavelength, employing nine subsystems sharing a primary system. The camera can capture the entire Earth's two-dimensional morphology and spectral fingerprints at a 10 km spatial resolution, with all spectral images acquired concurrently on a single detector. The camera's optical system is designed and simulated, and the stray light is analyzed and suppressed. Simulation and analysis results show that the camera can obtain high-quality images of the Earth's disk with a 2.5° field of view (FOV). The stray light is suppressed to less than 0.05% of the observed multispectral Earth radiation. The novel EAMC provides a new way to generate climate-relevant knowledge from the perspective of global Earth observation and has great potential for other applications in space-based remote sensing spectral imaging.

[Characteristics of Cadmium Concentration and Transport of Pueraria thornsonii in Farmland with Different Cadmium Pollution Degrees].

This study aimed to elucidate the cadmium (Cd) concentration and transport characteristics of Pueraria thornsonii in farmland with different Cd pollution degrees, so as to provide a reference basis for phytoremediation of Cd-contaminated farmland. The multi-point experiments in farmland with different Cd pollution degrees[ω(Cd) 0.32-38.08 mg·kg-1] were conducted, and the biomass (dry weight), Cd content, accumulation, concentration, and transport of Cd in P. thornsonii tissues under the main growing period were assessed. According to the results, for P. thornsonii, the tuber dry weight ranged from 5.04 to 11.98 t·hm-2, biomass ranged from 13.21 to 29.07 t·hm-2, and Cd accumulation ranged from 15.74 to 106.03 g·hm-2in the study area. The pattern of Cd uptake by P. thornsonii showed that the main vine>leaf>lateral branches>basal part of sti>tuber. The Cd content in P. thornsonii tissues considerably increased with soil Cd content (P<0.05), whereas the biomass decreased significantly (P<0.05). The Cd concentration and transport factor of aboveground parts in P. thornsonii showed a trend of initially falling, then increasing and decreasing again, whereas the Cd enrichment and transport coefficient of tubers gradually decreased. Correlation analysis revealed that the amount of Cd in the soil was a major predictor of Cd accumulation in P. thornsonii. Under light to moderate Cd contamination, the commercial portion of P. thornsonii (arrowroot)[ω(Cd) 0.03-0.22 mg·kg-1] was less than the standard limit for medicinal plants (≤ 0.30 mg·kg-1). In P. thornsonii from moderately contaminated areas, the Cd concentration and transport factor of aboveground parts were 2.43-7.97 and 3.02-9.81, respectively. This indicates that P. thornsonii is a prospective plant ideal for remediating Cd-contaminated soil because of its high capacity to transfer and enrich Cd.

The BRCA1/BARD1 complex recognizes pre-ribosomal RNA to facilitate homologous recombination.

The BRCA1/BARD1 complex plays a key role in the repair of DNA double-strand breaks (DSBs) in both somatic cells and germ cells. However, the underlying molecular mechanism by which this complex mediates DSB repair is not fully understood. Here, we examined the XY body of male germ cells, where DSBs are accumulated. We show that the recruitment of the BRCA1/BARD1 complex to the unsynapsed axis of the XY body is mediated by pre-ribosomal RNA (pre-rRNA). Similarly, the BRCA1/BARD1 complex associates with pre-rRNA in somatic cells, which not only forms nuclear foci in response to DSBs, but also targets the BRCA1/BARD1 complex to DSBs. The interactions between the BRCT domains of the BRCA1/BARD1 complex and pre-rRNA induce liquid-liquid phase separations, which may be the molecular basis of DSB-induced nuclear foci formation of the BRCA1/BARD1 complex. Moreover, cancer-associated mutations in the BRCT domains of BRCA1 and BARD1 abolish their interactions with pre-rRNA. Pre-rRNA also mediates BRCA1-dependent homologous recombination, and suppression of pre-rRNA biogenesis sensitizes cells to PARP inhibitor treatment. Collectively, this study reveals that pre-rRNA is a functional partner of the BRCA1/BARD1 complex in the DSB repair.

Orientated-quantitative computed tomography study on individualized axial safety target area of femoral neck screw channel and establishment of a stable spatial coordinate system based on anterior cortex of femoral neck basilar.

BACKGROUND: Frequent in-out-in femoral neck screws were reported potential huge iatrogenic-injury risks, related to axial safe target area (ASTA) of femoral neck screws channel. However, orientated-quantitative ASTA based on stable coordinate system was unreported before. METHODS: Three-dimensional reconstruction was performed on computed tomography (CT) images of 139 intact normal hips, and the intersection area, defined as ASTA, was obtained by superimposing the axial CT images of each femoral neck. Taking anterior cortex of femoral neck basilar (AC-FNB) as landmark, a coordinate system was established to measure the anterior-posterior diameter (D-AP), the superior-inferior diameter (D-SI) and the oblique angle respectively. Each intersection was overlaid up to the axial CT images to determine the coronal location of the ASTA boundaries. RESULTS: Each ASTA presented an inclined rounded triangle with a flat anterior base coincided with AC-FNB. There were significant sex differences in D-SI (male: 33.6±2.3 vs. female: 29.4±1.9 mm) and D-AP (male: 25.3±2.1 vs. 21.9±1.9 mm), P <0.001. D-SI was found to be positively correlated with D-AP ( R2 =0.6). All fluoroscopic visible border isthmus completely matched the corresponding ASTA boundaries. The oblique angle was 5-53° (male: 28.1±10.3°, female: 27.1±8.2°) without significant difference between sexes. CONCLUSION: The intersection method was employed to conveniently acquire orientated-quantitative individualized ASTA. Under this coordinate system, x-ray data of screws could be converted to axial coordinates in CT ASTA, which could help surgeons design combined screws configuration preoperatively and evaluate quantitatively their axial position intraoperatively.

CD36+ cancer-associated fibroblasts provide immunosuppressive microenvironment for hepatocellular carcinoma via secretion of macrophage migration inhibitory factor.

Hepatocellular carcinoma (HCC) is an immunotherapy-resistant malignancy characterized by high cellular heterogeneity. The diversity of cell types and the interplay between tumor and non-tumor cells remain to be clarified. Single cell RNA sequencing of human and mouse HCC tumors revealed heterogeneity of cancer-associated fibroblast (CAF). Cross-species analysis determined the prominent CD36+ CAFs exhibited high-level lipid metabolism and expression of macrophage migration inhibitory factor (MIF). Lineage-tracing assays showed CD36+CAFs were derived from hepatic stellate cells. Furthermore, CD36 mediated oxidized LDL uptake-dependent MIF expression via lipid peroxidation/p38/CEBPs axis in CD36+ CAFs, which recruited CD33+myeloid-derived suppressor cells (MDSCs) in MIF- and CD74-dependent manner. Co-implantation of CD36+ CAFs with HCC cells promotes HCC progression in vivo. Finally, CD36 inhibitor synergizes with anti-PD-1 immunotherapy by restoring antitumor T-cell responses in HCC. Our work underscores the importance of elucidating the function of specific CAF subset in understanding the interplay between the tumor microenvironment and immune system.

STRIPE3, encoding a human dNTPase SAMHD1 homolog, regulates chloroplast development in rice.

Chloroplast is an important organelle for photosynthesis and numerous essential metabolic processes, thus ensuring plant fitness or survival. Although many genes involved in chloroplast development have been identified, mechanisms underlying such development are not fully understood. Here, we isolated and characterized the stripe3 (st3) mutant which exhibited white-striped leaves with reduced chlorophyll content and abnormal chloroplast development during the seedling stage, but gradually produced nearly normal green leaves as it developed. Map-based cloning and transgenic tests demonstrated that a splicing mutation in ST3, encoding a human deoxynucleoside triphosphate triphosphohydrolase (dNTPase) SAMHD1 homolog, was responsible for st3 phenotypes. ST3 is highly expressed in the third leaf at three-leaf stage and expressed constitutively in root, stem, leaf, sheath, and panicle, and the encoded protein, OsSAMHD1, is localized to the cytoplasm. The st3 mutant showed more severe albino leaf phenotype under exogenous 1-mM dATP/dA, dCTP/dC, and dGTP/dG treatments compared with the control conditions, indicating that ST3 is involved in dNTP metabolism. This study reveals a gene associated with dNTP catabolism, and propose a model in which chloroplast development in rice is regulated by the dNTP pool, providing a potential application of these results to hybrid rice breeding.

Radiocarbon and genomic evidence for the survival of Equus Sussemionus until the late Holocene.

The exceptionally rich fossil record available for the equid family has provided textbook examples of macroevolutionary changes. Horses, asses, and zebras represent three extant subgenera of Equus lineage, while the Sussemionus subgenus is another remarkable Equus lineage ranging from North America to Ethiopia in the Pleistocene. We sequenced 26 archaeological specimens from Northern China in the Holocene that could be assigned morphologically and genetically to Equus ovodovi, a species representative of Sussemionus. We present the first high-quality complete genome of the Sussemionus lineage, which was sequenced to 13.4× depth of coverage. Radiocarbon dating demonstrates that this lineage survived until ~3500 years ago, despite continued demographic collapse during the Last Glacial Maximum and the great human expansion in East Asia. We also confirmed the Equus phylogenetic tree and found that Sussemionus diverged from the ancestor of non-caballine equids ~2.3-2.7 million years ago and possibly remained affected by secondary gene flow post-divergence. We found that the small genetic diversity, rather than enhanced inbreeding, limited the species' chances of survival. Our work adds to the growing literature illustrating how ancient DNA can inform on extinction dynamics and the long-term resilience of species surviving in cryptic population pockets.

Metal ratio mixing models clarify metal contamination sources to lake sediments in Yunnan, China.

Contaminated legacy sediments contribute to modern pollution loadings, particularly trace metals. These contributions are challenging to quantify as metal histories reconstructed from sediment records cannot be easily divided into legacy and concurrent contamination. In particular, the contribution from re-mobilization and delivery of legacy metals stored in catchment soil, colluvial, and fluvial environments are rarely considered or quantified when interpreting sediment records. Here, extended records of metals accumulation for a set of three lakes in Yunnan, China are compared with endmember chemistries using Monte Carlo-Markov Chain mixing models to help identify source contributions to the sediments. This approach allows attribution of metals transported by atmospheric and fluvial mechanisms in a region with a history of mining and metallurgy spanning millennia. These analyses reveal distinct source mixtures and demonstrate the sensitivity of lake records to basin sediment dynamics. In particular, substantial proportions of elevated metal concentrations in these lake systems seem to arise from soil contributions more than from atmospheric deposition of smelting emissions. The largest soil contributions seem to be in Erhai, a lake with erosion prone soils closely "connected" to the lake. Moreover, these invesigations illustrate the potential for mixing approaches to accommodate and clarify uncertainties in metal source and extraction as differences in extraction efficiency can be incorporated into source uncertainty estimates. Ultimately, these approaches emphasize the need to account for fluvial metal transport in interpretation of sediment histories.

Pre-ribosomal RNA reorganizes DNA damage repair factors in nucleus during meiotic prophase and DNA damage response.

In response to DNA double-strand breaks (DSBs), DNA damage repair factors are recruited to DNA lesions and form nuclear foci. However, the underlying molecular mechanism remains largely elusive. Here, by analyzing the localization of DSB repair factors in the XY body and DSB foci, we demonstrate that pre-ribosomal RNA (pre-rRNA) mediates the recruitment of DSB repair factors around DNA lesions. Pre-rRNA exists in the XY body, a DSB repair hub, during meiotic prophase, and colocalizes with DSB repair factors, such as MDC1, BRCA1 and TopBP1. Moreover, pre-rRNA-associated proteins and RNAs, such as ribosomal protein subunits, RNase MRP and snoRNAs, also localize in the XY body. Similar to those in the XY body, pre-rRNA and ribosomal proteins also localize at DSB foci and associate with DSB repair factors. RNA polymerase I inhibitor treatment that transiently suppresses transcription of rDNA but does not affect global protein translation abolishes foci formation of DSB repair factors as well as DSB repair. The FHA domain and PST repeats of MDC1 recognize pre-rRNA and mediate phase separation of DSB repair factors, which may be the molecular basis for the foci formation of DSB repair factors during DSB response.

Mitochondrial Phylogenomics of Tenthredinidae (Hymenoptera: Tenthredinoidea) Supports the Monophyly of Megabelesesinae as a Subfamily.

Tenthredinidae represents one of the external feeders of the most diverse superfamily, Tenthredinoidea, with diverse host plant utilization. In this study, four complete mitochondrial genomes (mitogenomes), those of Cladiucha punctata, Cladiucha magnoliae, Megabeleses magnoliae, and Megabeleses liriodendrovorax, are newly sequenced and comparatively analyzed with previously reported tenthredinid mitogenomes. The close investigation of mitogenomes and the phylogeny of Tenthredinidae leads us to the following conclusions: The subfamilial relationships and phylogenetic placements within Tenthredinidae are mostly found to be similar to the previously suggested phylogenies. However, the present phylogeny supports the monophyly of Megabelesesinae as a subfamily, with the sister-group placement of Cladiucha and Megabeleses outside of Allantinae. The occurrence of the same type of tRNA rearrangements (MQI and ANS1ERF) in the mitogenomes of Megabelesesinae species and the presence of apomorphic morphological characters also provide robust evidence for this new subfamily. The divergence and diversification times of the subfamilies appear to be directly related to colonization of the flowering plants following the Early Cretaceous. The origin time and diversification patterns of Megabelesesinae were also well matched with the divergence times of their host plants from Magnoliaceae.

Prognostic value of hemodynamic indices in patients with sepsis after fluid resuscitation.

BACKGROUND: Sepsis usually causes hemodynamic abnormalities. Hemodynamic index is one of the factors to identify the severity of sepsis and an important parameter to guide the procedure of fluid resuscitation. The present study investigated whether the assessment of hemodynamic indices can predict the outcomes of septic patients undergoing resuscitation therapy. AIM: To evaluate the prognostic value of hemodynamic indices in patients with sepsis after fluid resuscitation. METHODS: A retrospective study was conducted in 120 patients with sepsis at Hainan General Hospital/Hainan Affiliated Hospital of Hainan Medical University between October 2016 and October 2019. All patients were treated with sodium chloride combined with dextran glucose injection for fluid resuscitation. Patients' hemodynamic parameters were monitored, including heart rate (HR), cardiac index (CI), systemic vascular resistance index (SVRI), mean arterial pressure (MAP), central venous pressure (CVP), and central venous oxygen saturation. The prognostic value of hemodynamic indices was determined based on the prognosis status. RESULTS: During fluid resuscitation, 86 patients developed septic shock and 34 did not. Ninety-nine patients survived and 21 patients died at 28 d after the treatment. Heart rate, CI, mean arterial pressure, SVRI, and CVP were higher in patients with septic shock and patients who died from septic shock than in non-shock patients and patients who survived, and central venous oxygen saturation was lower in patients with shock and patients who died than in non-shock patients and the survivors (P < 0.05). When prognosis was considered as a dependent variable and hemodynamic parameters was considered as independent variables, the results of a logistic regression analysis showed that CI, SVRI, and CVP were independent risk factors for septic shock, and CI was an independent risk factor for 28-d mortality (P < 0.05). CONCLUSION: Hemodynamic indices can be used to evaluate the prognosis of septic patients after fluid resuscitation.

Contrasting ecosystem responses to climatic events and human activity revealed by a sedimentary record from Lake Yilong, southwestern China.

Global climate change and human activities have significantly impacted lake ecosystems at an accelerating rate in recent decades, but the differences between the responses of lake ecosystems to these two stressors remain unclear. Thus an improved understanding of the long-term influences of climatic and anthropogenic disturbances is necessary for the management of lake ecosystems. In order to address these issues, a sedimentary record was obtained from Lake Yilong in Yunnan Province in southwestern China, where the climate and natural environment are dominated by the Indian Summer Monsoon and there is a long history of human occupation and intensive human activity. The chronology is based on AMS 14C dates from 13 samples of plant macrofossils and charcoal, which show that the record spans the last ~12,000 yr. Geochemical indices were used to reconstruct hydro-climatic variations and lake ecosystem responses. The results indicate that a cold and humid climate prevailed from the late Pleistocene to the beginning of the Holocene, which was interrupted by an abrupt decrease in precipitation during 9.7-8.7 ka (1 ka = 1000 cal yr BP, corresponding to the 9.3 ka event). A persistent drying trend occurred during the middle and late Holocene, and there was an increase in the intensity of human activity during the past 1500 years. A comparison of the effects of a natural climatic event and human disturbance reveals contrasting lake ecosystem responses. The lake ecosystem was resilient to the 9.3 ka event and subsequently recovered; however, long-term human activity in the watershed, including deforestation and cultivation, reduced the stability of the lake ecosystem and positive feedback effects were strengthened, leading to the deviation of the system far from its previous stable state. It is concluded that, compared to climate change, human activities have had a much more serious impact on lake ecosystem.

Advanced treatment of secondary effluent organic matters (EfOM) from an industrial park wastewater treatment plant by Fenton oxidation combining with biological aerated filter.

This study investigated the advanced treatment of secondary effluent organic matters (EfOM) from an industrial park wastewater treatment plant (IPWTP) by Fenton oxidation process and its combination with biological aerated filter (BAF). The constituents of EfOM were characterized by using fluorescence excitation-emission matrix, and the results showed that the major components included aromatic proteins, soluble microbial products, humic and fulvic acid-like substances, and compounds associated with fluorescent region of Ex 250-300 nm/Em 600-700 nm. The EfOM was strongly resistant to biodegradation (biochemical oxygen demand (BOD5):chemical oxygen demand (COD) ratio at 0.11), resulting in less than 15% dissolved organic carbon (DOC) removal efficiency by the BAF reactor. The advanced treatment of EfOM by Fenton oxidation process led to maximum ~50% mineralization efficiency of EfOM under the optimal conditions of 2.0 mM FeII, 10 mM H2O2, pH 3.0 and 3.0 h of the reaction time. Particularly, Fenton oxidation treatment effectively improved the biodegradability of EfOM in the IPWTP secondary effluents, e.g., increasing the BOD5:COD ratio from 0.11 to 0.42. A synergistic combination of Fenton oxidation process with the BAF reactor offered desirable mineralization efficiencies of EfOM (>70%) at lower dosages of Fenton's reagents. The present results suggest that Fenton oxidation process combining with the BAF reactor can be a promising strategy for the advanced treatment of EfOM in IPWTP secondary effluents. This study provides guidance for the characterization and advanced treatment of EfOM in IPWTP secondary effluents for practical purpose.

Identification and biochemical characterisation of Acanthamoeba castellanii cysteine protease 3.

BACKGROUND: Acanthamoeba spp. are free-living amoeba that are ubiquitously distributed in the environment. This study examines pathogenic Acanthamoeba cysteine proteases (AcCPs) belonging to the cathepsin L-family and explores the mechanism of AcCP3 interaction with host cells. METHODS: Six AcCP genes were amplified by polymerase chain reaction (PCR). Quantitative real-time PCR was used to analyse the relative mRNA expression of AcCPs during the encystation process and between pre- and post-reactivated trophozoites. To further verify the role of AcCP3 in these processes, AcCP3 recombinant proteins were expressed in Escherichia coli, and the hydrolytic activity of AcCP3 was determined. The influence of the AcCP3 on the hydrolytic activity of trophozoites and the toxicity of trophozoites to human corneal epithelial cells (HCECs) was examined by inhibiting AcCP3 expression using siRNA. Furthermore, the levels of p-Raf and p-Erk were examined in HCECs following coculture with AcCP3 gene knockdown trophozoites by Western blotting. RESULTS: During encystation, five out of six AcCPs exhibited decreased expression, and only AcCP6 was substantially up-regulated at the mRNA level, indicating that most AcCPs were not directly correlated to encystation. Furthermore, six AcCPs exhibited increased expression level following trophozoite reactivation with HEp-2 cells, particularly AcCP3, indicating that these AcCPs might be virulent factors. After refolding of recombinant AcCP3 protein, the 27 kDa mature protein from the 34 kDa pro-protein hydrolysed host haemoglobin, collagen and albumin and showed high activity in an acidic environment. After AcCP3 knockdown, the hydrolytic activity of trophozoite crude protein against gelatin was decreased, suggesting that these trophozoites had decreased toxicity. Compared with untreated trophozoites or negative control siRNA-treated trophozoites, AcCP3-knockdown trophozoites were less able to penetrate and damage monolayers of HCECs. Western blot analysis showed that the activation levels of the Ras/Raf/Erk/p53 signalling pathways in HCECs decreased after inhibiting the expression of trophozoite AcCP3. CONCLUSIONS: AcCP6 was correlated to encystation. Furthermore, AcCP3 was a virulent factor in trophozoites and participated in the activation of the Ras/Raf/Erk/p53 signalling pathways of host cells.