The dense and toxic blooms formed by cyanobacteria in the aquatic environment have caused serious damage to public health and aquatic ecosystems. It is crucial to monitor and prevent cyanobacterial blooms in freshwater waters in a timely manner. Although the object detection method has been applied to the field of algae identification, there are some issues to be solved in the existing research. The main challenge is that the setting of the training set is too idealistic, which is separated from the actual water quality environment and cannot guide the actual algae identification and water quality monitoring. In this paper, we collect 2,024 algae microscopic images of algae from a reservoir in southern China and a data set for object detection. Aiming at the above problems, we proposed a morphable attention based algal tiny object detection strategy to sample and extract features of algae small targets more effectively. It is hoped that the model training and improvement results of this paper can help to establish a future algae real-time monitoring and water quality evaluation system.

Algal blooms have seriously affected the production and life of people and real-time detection of algae in water samples is a powerful measure to prevent algal blooms. The traditional manual detection of algae with a microscope is extremely time-consuming. In recent years, although there have been many studies using deep learning to classify and detect algae, most of them have focused on the relatively simple task of algal classification. In addition, some existing algal detection studies not only use small datasets containing limited algal species, but also only prove that object detection algorithms can be applied to algal detection tasks. These studies cannot implement the real-time detection of algae and timely warning of algal blooms. Therefore, this paper proposes an efficient self-organized detection system for algae. Benefiting from this system, we propose an interactive method to generate the algal detection dataset containing 28,329 images, 562,512 bounding boxes and 54 genera. Then, based on this dataset, we not only explore and compare the performance of 10 different versions of state-of-the-art object detection algorithms for algal detection, but also tune the detection system we built to its optimum state. In practical application, the system not only has good algal detection results, but also can complete the scanning, photographing and detection of a 2 cm × 2 cm, 0.1 mL algal slide specimen within five minutes (the resolution is 0.25886 μm/pixel); such a task requires a well-trained algal expert to work continuously for more than three hours. The efficient algal self-organized detection system we built makes it possible to detect algae in real time. In the future, with the help of IoT, we can use various smart sensors, actuators and intelligent controllers to achieve real-time collection and wireless transmission of algal data, use the efficient algal self-organized detection system we built to implement real-time algal detection and upload the detection results to the cloud to realize timely warning of algal blooms.

太湖流域地处长江下游,水系发达,是典型的平原水网地区,为明确空间和环境因子对太湖流域浮游植物功能类群分布的影响,本研究于2019年8-9月期间比较太湖流域包括河流、湖泊和水库的85个采样点的浮游植物、空间和环境特征.首次利用"人工智能图像识别技术",并结合传统人工镜检,共鉴定浮游植物250个分类单位,隶属于9门13纲28目57科110属,以绿藻门(45.6％)、硅藻门(19.2％)和蓝藻门(16.4％)为主.共划分功能类群31个,包括M、D、J、S1、C、Y、F、P、MP、H1、SN、W1、G、K和TB共15个优势功能类群.结果表明:1)太湖流域不同水体功能类群组成差异显著,差异主要来自M、J、S1、Y、P、H1类群;2)浮游植物功能类群分布具有一定的空间异质性,以太湖为中心向外形成3个聚类组,物种组成类型依次为蓝藻型、蓝藻—绿藻-硅藻型、绿藻-硅藻-蓝藻型,主要贡献类群依次为M、S1类群,M、S1、P、J类群,P、S1、J、D、C类群;3)在太湖流域,相对于环境因子而言,空间因子对浮游植物功能类群组成的影响更大.RDA分析表明,CODMn、NO3--N、TN、SD、pH值和DO与太湖流域浮游植物优势功能类群呈显著相关.过去十年,以控源截污为主导的环境管理政策已在太湖流域取得显著成效,但是对于以M、S1类群水华为特征的夏季太湖流域而言,蓝藻水华防控仍是必须面对的难题,有机污染物和氮的控制仍是关注重点,不同水体类型和浮游植物功能类群因地制宜的管控模式仍是关键.

Identifying chemicals with endocrine disrupting properties linked to disease outcomes is a key concern, as stated in the WHO-UNEP 2012 report on endocrine-disrupting chemicals. The chemical 9,9-bis[4-(2-hydroxyethoxy)phenyl]fluorene (BPEF) is widely and increasingly applied in synthesizing fluorene-based cardo polymers with superior optical, thermal and mechanical properties for various uses. However, little toxicological information is available regarding its safety. Here, we studied the endocrine disrupting property of BPEF by multiple toxicological tools and investigated its effects on female development in adolescent mice. Using the yeast two-hybrid bioassay, BPEF showed strong antiestrogenicity which was similar to that of tamoxifen, an effective antiestrogenic drug. In adolescent CD-1 mice, BPEF significantly decreased the uterine weight at relatively low doses and induced marked endometrial atrophy. Immunohistochemical staining and transcriptome analyses of the mice uteri revealed that BPEF could repressed the expressions of estrogen-responsive genes. Molecular simulation indicated that BPEF could be docked into the antagonist pocket of human estrogen receptor α, and the formation of hydrogen bonds and hydrophobic interactions between BPEF and the active site of receptor maintained their strong binding. All of the data demonstrated that BPEF possessed strong antiestrogenic property and might disrupt female development, suggesting it should be avoided in making products that might directly expose to people, particularly immature women.

藻类鉴定被广泛应用于藻类遗传学、生理学、生态学和应用藻类学,尤其是藻类调查和评估.然而,基于形态学的鉴定往往因为分类特征未出现或不典型、设备限制和人员经验欠缺等原因带来较大误差.随着测序技术的不断发展,分子标记已成为藻类鉴定的一个通用工具.由于藻类类群众多且差异很大,分子标记的选择成为藻类鉴定的关键.本文综述了蓝藻、硅藻、绿藻、甲藻、裸藻、隐藻、金藻、黄藻、红藻和褐藻等主要门类分子标记的选择及应用进展,包括分子标记选择原则、常用标记和相应序列数据库,以及各个分子标记在不同类群应用中的优缺点等.藻类分子鉴定源于编码核糖体RNA的基因(rDNA),发展于细胞核、线粒体、叶绿体DNA等.然而,当前藻类分子鉴定逐渐细化和完善,单一的核糖体DNA、内转录间隔区(ITS)和保守蛋白编码基因等短序列分子标记已经很难满足藻类鉴定的需求,多标记组合成为一种必然选择.同时,线粒体基因组、叶绿体基因组、核基因组、转录组和宏基因组等提供了更多遗传进化信息,弥补了短序列分子标记在系统分类应用中的不足.对于藻类鉴定,单纯依赖分子标记或形态学都不足以保证鉴定的准确性,采用将分子生物学、形态学、生理生化学等结合的多相学方法,才能准确地完成鉴定工作.此外,藻类分子数据库的建立和完善是未来分子鉴定的重要工作,快速鉴定方法也必将在未来获得广泛的应用和发展.

Fluorene-9-bisphenol (BHPF), a substitute for bisphenol A (BPA), has been widely used in the synthesis of polyester polymers. Studies have reported multiple BHPF toxicities but its effect on the liver remains unknown. In this study, we performed short-term and subchronic toxicity tests, as well as primary hepatocyte experiments, to investigate the hepatic toxicity of BHPF using CD-1 mice. And microarray was used to analyze the changes of global gene expression in the liver of mice treated with BHPF. The results showed that the liver coefficient and the activities of serum aminotransferases were obviously elevated by BHPF at doses of 27.8 mg/kg body weight (bw)/day or higher in mice treated for 10 days. Histological analysis showed obvious changes, including narrowed hepatic sinuses, dilated central vein, leucocyte infiltration, and cytoplasmic vacuolation, in the livers of mice treated with BHPF at dosages of 2 mg/kg bw/3-day and higher for 36 days. Microarray analyses revealed 2623 differentially expressed genes (DEGs) in the livers of mice treated with 50 mg/kg bw/day of BHPF for 3 days, which could be enriched in GO terms of T cell activation, leukocyte migration, and leukocyte chemotaxis and KEGG pathways of natural killer cell-mediated cytotoxicity and autoimmune thyroid disease. The top 10 hub DEGs, including LTF and MMP8, were observed in the protein-protein interaction network obtained via STRING database analysis, and are proposed as potential biomarkers for liver injury studies. Primary hepatocyte experiments demonstrated the hepatotoxicity of BHPF at concentrations of 10−6 M and higher. This study indicates that BHPF could cause liver injury at relatively low levels, suggesting that the risk of human BHPF exposure should be of concern.

Bisphenol A (BPA) is used in the production of plastic but has oestrogenic activity. Therefore, BPA substitutes, such as fluorene-9-bisphenol (BHPF), have been introduced for the production of so-called ‘BPA-free’ plastics. Here we show that BHPF is released from commercial ‘BPA-free’ plastic bottles into drinking water and has anti-oestrogenic effects in mice. We demonstrate that BHPF has anti-oestrogenic activity in vitro and, in an uterotrophic assay in mice, induces low uterine weight, atrophic endometria and causes adverse pregnancy outcomes, even at doses lower than those of BPA for which no observed adverse effect have been reported. Female mice given water containing BHPF released from plastic bottles, have detectable levels of BHPF in serum, low uterine weights and show decreased expressions of oestrogen-responsive genes. We also detect BHPF in the plasma of 7/100 individuals, who regularly drink water from plastic bottles. Our data suggest that BPA substitutes should be tested for anti-oestrogenic activity and call for further study of the toxicological effects of BHPF on human health.

The characteristics of tongue coating are very important symbols for disease diagnosis in traditional Chinese medicine (TCM) theory. As a habitat of oral microbiota, bacteria on the tongue dorsum have been proved to be the cause of many oral diseases. The high-throughput next-generation sequencing (NGS) platforms have been widely applied in the analysis of bacterial 16S rRNA gene. We developed a methodology based on genus-specific multiprimer amplification and ligation-based sequencing for microbiota analysis. In order to validate the efficiency of the approach, we thoroughly analyzed six tongue coating samples from lung cancer patients with different TCM types, and more than 600 genera of bacteria were detected by this platform. The results showed that ligation-based parallel sequencing combined with enzyme digestion and multiamplification could expand the effective length of sequencing reads and could be applied in the microbiota analysis.

The estrogenicity of parabens at human exposure levels has become a focus of concern due to the debate over whether the estrogenicity of parabens is strong enough to play a role in the increased incidence of breast cancer. In this study, the uterotrophic activities of methylparaben (MP) and ethylparaben (EP) at doses close to the acceptable daily intake as allocated by JECFA were demonstrated in immature Sprague-Dawley rats by intragastric administration, and up-regulations of estrogen-responsive biomarker genes were found in uteri of the rats by quantitative real-time RT–PCR (Q-RT-PCR). At the same time, the urinary concentrations of MP and EP, as measured by gas chromatography–mass spectrometry (GC-MS) in rats that received the same doses of MP and EP, were found to be near the high urinary levels reported in human populations in recent years. These results show the in vivo estrogenicity of MP and EP at human exposure levels, and indicate that populations exposed to large amounts of MP and EP may have a high burden of estrogenicity-related diseases. In addition, a molecular docking simulation showed interaction between the parabens and the agonist-binding pocket of human estrogen receptor α (hERα).

Background: 16S rRNA genes have been widely used for phylogenetic reconstruction and the quantification of microbial diversity through the application of next-generation sequencing technology. However, long-read sequencing is still costly, while short-read sequencing carries less information for complex microbial community profiling; therefore, the applications of high throughput sequencing platforms still remain challenging in microbial community reconstruction analysis.Results: Here, we developed a method to investigate the profile of aligned 16S rRNA gene sequences and to measure the proper region for microbial community reconstruction, as a step in creating a more efficient way to detect microorganism at the genus level. Finally, we found that each genus has its own preferential genus-specific amplicons for a genus assignment, which are not always located in hyper variable regions (HVRs). It was also noted that the rare genera should contribute less than dominant ones to the common profile of the aligned 16S rRNA sequences and have lower affinity to the common universal primer.Conclusions: Therefore, using multiple 16S rRNA regions rather than one "universal" region can significantly improve the ability of microbial community reconstruction. In addition, we found that a short fragment is suitable for most genera identifications, and the proper conserved regions used for primer design are larger than before. (C) 2016 Elsevier Ltd. All rights reserved.

To assess the impact of sanitation of a living environment on gut microbiota and development of the immune system, we raised BALB/c mice under three distinct environmental conditions: a specific pathogen-free animal room (SPF), a general animal room (XZ) and a farmhouse (JD). All other variables like diet, age, genetic background, physiological status and original gut microbiota were controlled for in the three groups. Using high-throughput sequencing of the 16S rRNA gene, we found that each mouse group had a specific structure of the gut microbial community. Groups JD and XZ harboured a significantly more diverse and richer gut microbiota than did group SPF. Bacteroidetes were significantly more abundant in groups XZ and JD than in group SPF, whereas Firmicutes showed the inverse pattern. Total serum immunoglobulin E (IgE) levels were significantly lower in groups XZ and JD than in group SPF. There were no significant differences in gut microbiota diversity and serum IgE concentration between groups JD and XZ, but we found higher abundance of dominant genera in the gut microflora of group JD. We conclude that exposure to soil, house dust and decaying plant material enhances gut microbial diversity and innate immunity. Our results seem to provide new evidence supporting the hygiene hypothesis.

Benzylparaben (BzP), a type of parabens being used as a preservative agent in cosmetics, food, and pharmaceutical products, may be ingested by humans. In this study, we performed an immature uterotrophic assay using Sprague Dawley (SD) rats by intragastric administration to determine the estrogenic effects of BzP and found significant increases in uterine weight with doses of 0.16mg/kg body weight and higher (P<0.05). The in vivo estrogenicity of BzP was supported by in vitro results from the human estrogen receptor α (hERα)-coactivator recruiting assay and in silico molecular docking analysis performed in this study. The in vitro estrogenic activity of BzP can be observed at concentrations of 1.0×10−8 M and higher. Molecular docking analysis showed that BzP fits well into the agonist pocket of hERα. The lowest observed effect dose (LOED) (0.16mg/kg/day) of BzP is much lower than the documented LOEDs of other parabens. Actual risk may exist for people who consume a diet high in BzP or use BzP-laden cosmetics. In addition, we tested the sensitivity of Wistar rats to 17β-estradiol by immature uterotrophic assay, and no obvious uterotrophic response was observed in the rats given doses up to 100μg/kg body weight.

Parabens are p-hydroxybenzoic acid esters that have been used extensively as preservatives in foods, cosmetics, drugs and toiletries. These intact esters are commonly detected in human breast cancer tissues and other human samples, thus arousing concern about the involvement of parabens in human breast cancer. In this study, an in vitro nuclear receptor coactivator recruiting assay was developed and used to evaluate the binding activities of parabens, salicylates and benzoates via antagonist competitive binding on the human oestrogen-related receptor γ (ERRγ), which is known as both a diagnostic biomarker and a treatment target of breast cancer. The results showed that all of the test parabens (methyl-, ethyl-, propyl-, butyl- and benzylparaben) possessed clear inverse antagonist activities on ERRγ, with a lowest observed effect level (LOEL) of 10−7M and the 50% relative effective concentrations (REC50) varying from 3.09×10−7 to 5.88×10−7M, whereas the salicylates possessed much lower activities and the benzoates showed no obvious activity. In silico molecular docking analyses showed that parabens fitted well into the active site of ERRγ, with hydrogen bonds forming between the p-hydroxyl group of parabens and the Glu275/Arg316 of ERRγ. As the paraben levels reported in breast cancer tissues are commonly higher than the LOELs observed in this study, parabens may play some role via ERRγ in the carcinogenesis of human breast cancer. In addition, parabens may have significant effects on breast cancer patients who are taking tamoxifen, as ERRγ is regarded as a treatment target for tamoxifen.

Background: Natural or endogenous sense/antisense miRNAs, located on sense and antisense strands in the same genomic region, respectively, are detected recently. However, little is known about these miRNA pairs, especially for their distributions in different animal species. We herein present systematic analysis of them in human, mouse and rat miRNAs, and their expression patterns based on deep sequencing datasets.Methods and results: The phenomenon of miRNA-miRNA interaction could be detected in different animal species. The common miRNAs pairs were found across species. These miRNA pairs could form miRNA: miRNA duplex with complete complementary structure, and were prone to be located on specific chromosomes. They might be homologous miRNA genes (especially in human), or clustered in a gene cluster (especially in rat), or simultaneously detected in different genomic regions due to multicopy pre-miRNAs. Remarkably, some miRNA pairs, located in different genomic regions, also showed complementarity as well as endogenous sense/antisense miRNAs. Based on published deep sequencing datasets, one member of miRNA pairs always was abundantly expressed, whereas another was quite rare. Rare common target mRNAs of these miRNA pairs were predicted.Conclusions: Interaction between miRNAs and significant expression divergence implied complex potential mutual regulatory pattern in the miRNA world. The study would enrich miRNA regulatory network. (C) 2012 Elsevier B.V. All rights reserved.

BACKGROUND:The multiplexing becomes the major limitation of the next-generation sequencing (NGS) in application to low complexity samples. Physical space segregation allows limited multiplexing, while the existing barcode approach only permits simultaneously analysis of up to several dozen samples.RESULTS:Here we introduce pair-barcode sequencing (PBS), an economic and flexible barcoding technique that permits parallel analysis of large-scale multiplexed samples. In two pilot runs using SOLiD sequencer (Applied Biosystems Inc.), 32 independent pair-barcoded miRNA libraries were simultaneously discovered by the combination of 4 unique forward barcodes and 8 unique reverse barcodes. Over 174,000,000 reads were generated and about 64% of them are assigned to both of the barcodes. After mapping all reads to pre-miRNAs in miRBase, different miRNA expression patterns are captured from the two clinical groups. The strong correlation using different barcode pairs and the high consistency of miRNA expression in two independent runs demonstrates that PBS approach is valid.CONCLUSIONS:By employing PBS approach in NGS, large-scale multiplexed pooled samples could be practically analyzed in parallel so that high-throughput sequencing economically meets the requirements of samples which are low sequencing throughput demand.

Salicylate esters (SEs), a class of chemicals extensively used as flavor and fragrance additives in foods, beverages and a wide variety of consumer products, are suspected to have estrogenic activity based on chemical analysis of in silica molecular docking. We evaluated the estrogenic potentials of phenyl salicylate (PhS), benzyl salicylate (BzS), phenethyl salicylate (PES), ethyl salicylate (ES) and methyl salicylate (MS) using an in vitro human estrogen receptor α (hERα)-coactivator recruiting assay and in vivo immature rodent uterotrophic bioassays. We found that PhS, BzS and PES showed obvious in vitro hERα agonistic activities; BzS in particular exhibited a higher estrogenic activity compared to bisphenol A (BPA). The uterine weights were significantly increased in mice treated with 11.1, 33.3, 100 and 300 mg/kg/day BzS and 33.3mg/kg/day PES and rats treated with 3.7, 11.1, 33.3 and 100mg/kg/day BzS for 3 days (P<0.05). Finally, we transformed the daily intakes and the dermal exposures of SEs in the real world into estradiol equivalent concentrations (EEQs). We found that the EEQ of BzS daily intake in consumers in the U.S. and the EEQs of dermal BzS and PES exposure among high-volume users worldwide were higher than the maximum secure daily estradiol intake recommended by the U.S. Food and Drug Administration (FDA). In particular, the EEQ for dermal BzS exposure was up to 162 ng EEQ/kg, which is 3.3 times higher than the maximal acceptable daily E(2) intake recommended by the Joint FAO/WHO Expert Committee on Food Additives (JECFA).

Previously we have developed polyacrylamide gel-based DNA microarray to genotype single nucleotide polymorphism (SNP) in a large number of samples, which has been proved as a simple and robust platform for high-throughput SNP screening. Here we improved this method to detect multiple SNPs by introducing multiplex polymerase chain reaction (multiplex-PCR) and immobilizing the products of multiplex-PCR to fabricate gel-based microarray and applying universal dual-color detectors in hybridization. In this report, five SNPs (rs191296, rs2280073, rs17599165, rs17599416 and rs7660336) of GABRA4 gene were chosen and successfully analyzed with the improved platform. Our experiment demonstrated that 3-dimentional polyacrylamide gel-based microarray of multiplex-PCR products make the platform for multiple SNPs genotyping becoming more labor-saving and time-saving. Appling the universal dual-color fluorescent detectors can reduce the cost over two-thirds for multiple SNPs analysis. It is concluded that the multiplex-PCR combined with the gel-based microarray hybridized with universal dual-color fluorescent detectors is efficient, rapid and simple for the detection of a single nucleotide mismatch, and may be very competitive in the efficiency, fidelity and cost for constructing DNA microarrays.

Objective: PCR-DGGE (denaturing gradient gel electrophoresis) was applied to analyze autism gut microbial communities. Materials and methods: We extracted the total microbial genome DNA of each fecal sample from two group subjects: the control and ASD (autism spectrum disorders) groups, amplified the V3 regions of microbial 16S rDNA genes. The gut microbial diversities were detected by PCR- DGGE. Results: DGGE analysis revealed diverse bacterial populations among the individuals and between the two groups, the similarity values of the comparisons of the bacterial DGGE bands patterns of the control and ASD groups ranged from 0.41 to 0.72 and 0.20 to 0.48 respectively. Conclusions: PCR-based DGGE can be used to detect the intestinal microbial diversities as an effective tool, and the intestinal microbial diversities may take a significant role in the autistic etiology.

OBJECTIVE:To genotype single nucleotide polymorphisms (SNPs) in a large number of samples by applying three-dimensional polyacrylamide gel-based microarray.METHODS:The method relies on copolymerization of acrylamide-modified PCR products with acrylamide monomers and acryl-modified slides to prepare gel-based microarray. Then array is hybridized with a pair of specific probes and the two universal dual-color fluorescent detectors labeled with Cy3 or Cy5 respectively (Tag1 and Tag2). Electrophoresis is used in post-hybridization to remove the nonspecifically bound targets and mismatches. Finally, genotyping is based on the images captured through two-color fluorescent scanning.RESULTS:The 3-D gel-immobilization of nucleic acids has a high immobilization yield and good hybridization efficiency. As universal dual-color fluorescent detectors are used, it is not required that specific probes be labeled for all SNPs, therefore the expense for synthesis can be reduced considerably. Electrophoresis in post-hybridization can enhance the capability for discriminating a single nucleotide mismatch from the perfectly matched sequence and improve the signal-to-noise ratio significantly.CONCLUSION:The gel-based microarray is a rapid, simple and high-throughput method for SNPs genotyping and may be very competitive in the efficiency, fidelity and cost for constructing DNA microarrays, which will hold significant promise for applications in human DNA diagnostics.

Single nucleotide polymorphisms (SNPs) are important markers which can be used in association studies searching for susceptible genes of complex diseases. High-throughput methods are needed for SNP genotyping in a large number of samples. In this study, we applied polyacrylamide gel-based microarray combined with dual-color hybridization for association study of four BDNF polymorphisms with autism. All the SNPs in both patients and controls could be analyzed quickly and correctly. Among four SNPs, only C270T polymorphism showed significant differences in the frequency of the allele (χ2 = 7.809, p = 0.005) and genotype (χ2 = 7.800, p = 0.020). In the haplotype association analysis, there was significant difference in global haplotype distribution between the groups (χ2 = 28.19，p = 3.44e-005). We suggest that BDNF has a possible role in the pathogenesis of autism. The study also show that the polyacrylamide gel-based microarray combined with dual-color hybridization is a rapid, simple and high-throughput method for SNPs genotyping, and can be used for association study of susceptible gene with disorders in large samples.