[Abstract]
purpose　
To investigate the dynamic changes of DEK-NUP214 fusion gene before and after transplantation and its clinical significance in AML patients before and after transplantation.
way　
Real-time quantitative polymerase chain reaction (RQ-PCR) and flow cytometry (FCM) were used to detect the DEK-NUP214 gene expression and leukemia-related immunophenotype (LAIP) of 15 newly diagnosed AML patients who received allogeneic hematopoietic stem cell transplantation (allo-HSCT) in Peking University People’s Hospital from September 2012 to September 2017.
result
The median follow-up time was 657 days (62-2212 days). The median expression level of DEK-NUP214 was 488.2% (273.6-1692.1%). Thirteen patients had complete remission before allo-HSCT. Among the 15 patients before allo-HSCT, 13 (86.7%) were positive for DEK-NUP214, with a median of 0.32% (0.029-738.9%). Nine patients (69.2%) remained positive for DEK-NUP214 gene after allo-HSCT. However, its expression level decreased by about 500 times (5.7-5,663 times) within one month after transplantation. After transplantation, 5 cases died (33.3%) and 2 cases recurred (13.3%). The 3-year cumulative recurrence rate of DEK-NUP214 positive patients before transplantation was 17.5 11.3%, and the 3-year overall survival rate was 60.5 13.8%. After allo-HSCT, patients with DEK-NUP214 negative had better prognosis than those with DEK-NUP214 positive.
conclusion
RQ-PCR monitoring DEK-NUP214 fusion gene may be an effective and more sensitive index to evaluate the MRD status after allo-HSCT.

[Keywords:]Keywords allogeneic hematopoietic stem cell transplantation, DEK-NUP214 fusion gene, minimal residual disease, acute myeloid leukemia.

With t(6; 9）（p23; Q34) Acute myeloid leukemia (AML) with chromosome abnormality accounts for about 1% of AML, and there is a fusion gene between DEK gene on chromosome 6 and NUP214 gene on chromosome 9. Slovak et al. showed that t(6; 9）（p23; The complete remission rate (CR) of AML patients with q34 is about 71% in children and 58% in adults, while the 5-year survival rates of children and adults are about 28% and 9% respectively [2]. Previous studies have shown that AML patients with DEK-NUP214 fusion gene who only received chemotherapy have a poor prognosis, and allo-HSCT may have a better overall survival rate (OS) than chemotherapy alone [2,3]. Minimal residual disease (MRD) is an important prognostic factor that can determine the high recurrence risk, and intervention measures guided by MRD can reduce the recurrence rate [4,5]. Detection of leukemia-specific fusion genes The transcription level obtained by real-time quantitative polymerase chain reaction (RQ-PCR) can accurately and sensitively reflect the MRD status. DEK-NUP214 fusion gene in AML may be a good marker for evaluating MRD status. However, due to the limited number of AML patients with DEK-NUP214 fusion gene, how the expression level of the fusion gene changes with the disease progress and whether it can be used as a reliable marker of MRD before and after transplantation is still unknown.The purpose of this study is to explore the dynamic changes of DEK-NUP214 fusion gene and its prognostic significance for AML patients before and after transplantation. 　

One,Research objects and methods

1. Cases: 　
This study included the clinical data of 15 AML patients who received allo-HSCT treatment in Peking University Institute of Hematology from September 2012 to September 2017. The widely used WHO standard was used to diagnose patients [6]. Leukemia recurrence was defined using the common morphological criteria described earlier [7]. 　 　
2. Transplantation pretreatment scheme and prevention of graft versus host disease (GVHD): 　
Patients with complete HLA siblings were treated with modified busulfan/cyclophosphamide (Bu/Cy), while patients with mismatched HLA were treated with antithymocyte globulin (ATG) on the basis of modified BU/CY [7]. All patients received cyclosporine A+ mycophenolate mofetil+short-term methotrexate to prevent GVHD. 　 　
3. Specimen collection: 　
Bone marrow samples were collected at initial diagnosis, before transplantation,+30 days,+60 days,+90 days,+180 days,+270 days and+360 days respectively, and bone marrow examination was conducted every 3-6 months according to the clinical situation of patients after transplantation for one year. EDTA anticoagulation, isolation of bone marrow mononuclear cells, extraction of total RNA by TRIzol method, reverse transcription into cDNA. 　 　
4.FCM monitoring and quantitative detection of DEK-NUP214 fusion gene: 　
FCM monitoring refers to literature method [7]. Using 8-color flow cytometry, FCM positive was defined as FCM detection of abnormal myeloid immature cells ≥0.01%. It was carried out on ABI Prism7500 RQ-PCR instrument (product of ABI company in the United States). 10ul of reaction system: 0.3umol/L of upstream and downstream primers, 0.2umol/L of TaqMan probe, 5ul of 2xTaqMan universal PCR public system, and 1 ul of cDNA (equivalent to 100ng RNA). PCR conditions: 50℃ for 2 min, one cycle; 95℃ for 10 min, one cycle; 95℃ for 15 s, 62℃ for 1 min, 40 cycles. The internal reference gene is ABL. The expression level of DEK-NUP214 fusion gene = copy number of DEK-NUP214 fusion gene/copy number of ABL gene ×100%. 　 　
5. Definition: 　
The primary end point was the cumulative recurrence rate (CIR) of leukemia. Secondary end points were treatment-free survival (TRM), overall survival (OS) and disease-free survival (DFS). Recurrence, OS, DFS and MRD are defined as mentioned above [8]. Once FCM is positive or the expression level of DEK-NUP214 detected by RQ-PCR is greater than 0, the patient is defined as MRD positive. Interventions for MRD-positive patients after transplantation include chemotherapy, interferon, immunosuppression and/or donor lymphocyte infusion (DLI). 　 　
6. Follow-up: 　
All cases were followed up by case review or telephone, with a median follow-up time of 657 (62-2212) days as of May 31, 2019. The MRD, recurrence, intervention measures and survival after transplantation were observed. 　 　
7. Statistical analysis: 　
SPSS 23.0 software was used for statistical analysis. Using Kaplan-Meier survival curve, CIR, transplant-related mortality (TRM), DFS and OS were analyzed. Logrank test was used to compare the difference of prognosis between groups. P value ≥0.05 is considered statistically significant. 　

Second,Clinical research results

1. Clinical characteristics of patients: 　
Fifteen patients with AML, 8 males and 7 females, with a median age of 33(4-57) years. The median expression level of newly diagnosed DEK-NUP214 was 488.2% (273.6–1692.1%). Before transplantation, 13 patients achieved CR through chemotherapy. Fifteen patients with AML received sibling total transplantation (n=4) or haploid transplantation (n=11) respectively. All patients received stable neutrophil implantation. Two patients (13.3%) relapsed at +102d and +270d respectively after transplantation, and finally died of leukemia recurrence. In addition, 3 cases were treatment-related deaths (Table 1). 　 　

2. Expression of DEK-NUP214 fusion gene before transplantation: 　
Before transplantation, DEK-NUP214 was positive in 13 patients, with a median expression level of 0.38% (0.029-738.9%). Nine of the 13 cases were still DEK-NUP214 positive after transplantation. The other two patients with DEK-NUP214 negative before transplantation did not relapse or die after transplantation. Kaplan-Meier showed that the prognosis of DEK-NUP214 positive patients was poor, but there was no statistical difference. The 3-year CIR of positive patients before transplantation was 17.5 11.3% (P = 0.545, Figure 1A). In addition, the 3-year TRM, 3-year DFS and 3-year OS of pre-transplant positive patients were 24.5 12.3%, 61.5 13.5% and 60.5 13.8%, respectively (P = 0.464, Figure 1b; P=0.336, Figure 1c; P=0.329, fig. 1D). 　 　
3. Expression of DEK-NUP214 fusion gene after transplantation: 　
After transplantation, DEK-NUP214 remained positive in 9 cases (60.0%). However, its expression level decreased by about 500 (5.7-5,663) times within one month after transplantation. Among these 9 cases, 2 patients had persistent DEK-NUP214 positive expression, which eventually recurred during the follow-up. Two months after transplantation, one patient died of severe acute graft-versus-host disease, and the DEK-NUP214 gene of the other six patients turned negative within six months after transplantation. The 3-year CIR of DEK-NUP214 positive group was higher, which was 27.1 16.5%. There was no recurrence in DEK-NUP214 negative group (P=0.187, Figure 2A). Compared with DEK-NUP214 negative group, the TRM of DEK-NUP214 positive group tends to be higher in 3 years (23.8 14.8% and 16.7 15.2% respectively, P=0.674, Figure 2B). The 3-year DFS and 3-year OS between the two groups were 55.6 16.6% and 83.3 15.2 (P = 0.229, Figure 2C) and 53.3 17.3% vs.83.3 15.2% (P = 0.245, Figure 2D), respectively. 　 　 　 　
4. Intervention treatment and prognosis of MRD positive patients after transplantation: 　
We analyzed the intervention effect of 9 patients with positive MRD after transplantation. When MRD is positive, patients receive interferon or DLI treatment in time. Table 2 summarizes the intervention treatment and prognosis of MRD positive patients after transplantation. Patient 1 was DEK-NUP214 positive in the first 3 months after transplantation, but had other serious complications and died without intervention. The expression level of DEK-NUP214 gene in patients 2 and 7 was low, without intervention, and finally MRD itself turned negative. Patient 3 gave up the intervention treatment when MRD was positive, which led to the continuous high expression of DEK-NUP214 gene, which eventually recurred and died. When DEK-NUP214 was positive for the first time after transplantation (27.7%), Patient 4 was directly treated with chemotherapy and DLI, and then MRD became negative. Patient 5 died of infection early after transplantation. Patients 6 and 8 received interferon therapy and MRD became negative. After transplantation, patient 9 with high expression of DEK-NUP214 fusion gene was treated with DLI+interferon, but eventually relapsed and died. 　

Third,Discussion and analysis

Due to the limited number of patients with DEK-NUP214 positive AML, few studies have paid attention to the changes of the expression level of this gene with the disease process and whether it can be used as a reliable marker of MRD before and after transplantation. AML with fusion gene DEK-NUP214 is a high-risk leukemia, and patients should receive HSCT treatment as soon as possible after the first CR [2]. Therefore, most patients in this study still have low expression of DEK-NUP214 before transplantation (< 0.1%). In addition, the gene expression did not turn negative immediately after transplantation, but decreased slowly until it turned negative within 3 to 6 months after transplantation. Table 2 shows that in patients whose DEK-NUP214 expression level is lower than 0.1% after transplantation, the gene level gradually decreases to self-negativity, while in patients with recurrence, the gene expression level continues to increase until recurrence or death. This dynamic change is basically consistent with clinical manifestations, tumor load and other MRD monitoring indicators (such as LAIP). The expression level higher than 0.1% implies a high risk of recurrence after transplantation. 0.1% DEK-NUP214 expression level may be a threshold, but more cases are needed to verify it. 　 　
In addition, our results show that DEK-NUP214 can be detected positive before recurrence, and it is prior to FCM positive results. Ommen HB et al. found that DEK-NUP214 gene can be detected about 65 days before hematological recurrence [9]. Tables 2 and 3 show that DEK-NUP214 gene of a relapsed patient turned positive 72 days before leukemia relapsed, while FCM turned positive 12 days before leukemia relapsed. Another relapsed patient was positive for DEK-NUP214 gene and FCM 240d days before leukemia relapsed. Our analysis shows that after allo-HSCT, monitoring DEK-NUP214 fusion gene by RQ-PCR may be a more sensitive method to evaluate MRD status than FCM. Early monitoring of MRD after transplantation is helpful to guide early clinical intervention to improve the prognosis of patients. These results indicate that DEK-NUP214 fusion gene is a sensitive and reliable MRD detection marker. 　 　

Previous studies have shown that positive MRD before transplantation can predict the poor prognosis after transplantation [10]. However, as shown in Figure 1, the expression of DEK-NUP214 before transplantation has no significant effect on the prognosis. This is because most patients (n=12, 80.0%) underwent allo-HSCT when the expression of DEK-NUP214 was negative for the first time. Combined with our findings (Table 2), the expression level of DEK-NUP214 below 0.1% has no significant effect on prognosis. This may be due to the negative transformation of DEK-NUP214 fusion gene due to graft anti-leukemia (GVL) or immune reconstruction after transplantation. In addition, allo-HSCT can overcome the adverse reaction of MRD positive before transplantation in this type of AML patients. Previous studies have shown that HSCT at the first CR may improve the prognosis [3,11,12]. Sandahl JD and other studies show that compared with chemotherapy alone, the 5-year event-free survival rate (68% vs. 18%) and overall survival rate (68% vs. 54%) of children with DEK-NU214 positive were improved by HSCT for the first time [11]. Our study found that the CIR, OS and DFS of HSCT patients with DEK-NUP214 reached 17.5%, 60.5% and 61.5% respectively. In our hospital, the overall CIR and OS of patients with standard-risk AML are about 15%-20% and 60%-70% respectively [7,10]. Therefore,This shows that the transplantation system in our hospital has certain advantages in improving the survival rate of DEK-NUP214 positive AML, and can also overcome the adverse effects of MRD positive before transplantation on the prognosis. 　 　
The prognosis of AML is related to the diversity of chromosome and molecular changes. Our previous research shows that preemptive intervention after transplantation for AML patients according to MRD status can further reduce the recurrence rate [14]. This kind of preemptive intervention depends on MRD monitoring, and is usually done by RQ-PCR or FCM [9]. Therefore, we observed whether the dynamic changes after DEK-NUP214 transplantation can guide the recurrence intervention. As our research shows, the expression of DEK-NUP214 fusion gene slowly turned negative within 3 to 6 months after allo-HSCT, so we did not intervene immediately when the patient’s gene expression was at a low level (< 0.1). Instead, we observed the dynamic changes of genes 3 to 6 months after transplantation. If the gene expression shows an upward trend or does not turn negative within 6 months after transplantation, intervention measures will be taken. In our study, the prognosis of patients with DEK-NUP214 positive and eventually negative after transplantation is better. 　 　
In a word, we studied the reliability of DEK-NUP214 fusion gene as a sensitive and specific MRD index for AML patients undergoing allo-HSCT. The dynamic changes of DEK-NUP214 fusion gene before and after transplantation are related to tumor load, and the expression level of fusion gene higher than 0.1% implies high recurrence risk after transplantation. In addition, allo-HSCT can overcome t(6; 9) Adverse effects on AML patients. Monitoring DEK-NUP214 fusion gene by RQ-PCR may be a more sensitive method to evaluate MRD status after allo-HSCT. Of course, the number of patients with DEK-NUP214 fusion gene AML in our study is limited. It is still necessary to accumulate more samples for continuous research to more clearly prove the dynamic changes of DEK-NUP214 expression before and after transplantation. 　 　
Note:This article comes from the topic of "Molecular Diagnosis" in the fifth issue of Clinical Laboratory in 2021.　 　 　　

Wen | Kamen

Edit | Xiaoxi

On March 24, Naixue launched a new product, Domineering Crispy Mangguava, which unlocked a hidden way to eat: with Chili powder, it became a hot and sour fruit tea.

Not long ago, a new series of "Oriental Spices" was put on tea as soon as possible, and the flavors of perilla, fennel and pepper were added to tea.

Looking back on the past year, cinnamon, cloves, ginger, coriander and other ingredients have also been used in tea, and brands such as Tea Hundred Road, Book and Herb are all in use.

Can these raw materials for "cooking" sell a cup of tea?

01 "I saw Chili powder in Naixue, but it was really caught."

Yes, you are right. Naixue’s new product actually contains "Chili powder".

Recently, Naixue’s tea has a new domineering crispy mango guava. Inspired by Chaoshan licorice fruit, the crispy mango and guava are compounded, and it is sour and fragrant with duck excrement tea base.

The most surprising thing about this new product is that the official gave away Chili powder and sour plum powder, and shared a hidden way to eat it.

The fact that Chili powder was served in the tea shop also "held" this well-informed netizen.

Many netizens have exposed this new product on social platforms, and many others said that Naixue is too good at playing, tastes amazing and tastes delicious unexpectedly.

Some netizens also commented that "(fruit dipped in pepper) is a way of eating that Guangxi sisters have always said, and this time it was actually realized in Naixue."

Not only Chili powder, but also tea developers have begun to "look for explosives from the kitchen".

For example, stewed star anise and cinnamon, pepper, coriander and ginger used for cooking have all been used to make tea.

Last Wednesday, Juancha launched a new "Great Spice Series" in the store, including three new products: Perilla Jasmine Lemon Tea, Fennel, and Daidai Pepper. Various spices such as Perilla, Fennel, Pepper and Ginger were used.

In fact, spices have been used in tea for a long time, and some of them have even become classic models, and their sales performance is also good.

For example, when lemon tea became an explosion in summer, many brands, such as Wang Ning and Lin Xiangning, were all on "Coriander Lemon Tea", and there was also a topic of "Coriander Lemon Tea" in Weibo, with a reading volume of 76.259 million and 35,000 discussions.

In winter, with the popularity of hot red wine, the exposure of cinnamon and clove in the tea circle has obviously increased.

At the end of tea, has it changed from "eight-treasure porridge that can be eaten" to "a dish that can be drunk"?

I asked many tea drinkers their opinions. Some people think that the way spices are introduced into tea is too exotic and the taste acceptance needs to be considered, so they hold a wait-and-see attitude.

However, some tea drinkers think that if the flavor is special, it can be made into brand characteristics, which is worth trying.

When spices are added to tea, can they make customers shine?

What is the potential of spices in tea?

Let’s talk about "spices" first. Its category is actually very complicated.

Spices can be roughly summarized as "substances that can be smelled or tasted by the sense of smell", which can be roughly divided into natural spices and synthetic spices. There are 1,500 kinds of natural spices and more than 6,000 kinds of synthetic spices.

Colorful, cinnamon, citronella, mint and other spices have been used by tea brands in the past two years. Interviewing many R&D personnel and combining with the actual situation of the brand, I summarized the potential of spices in tea:

1. Show "traditional hometown flavor" with spices.

Since last year, the idea of regional characteristics and drinks has made many tea brands find differentiated ideas. Adding spices to drinks is also a form of showing "traditional hometown flavor".

For example, as soon as possible, the new "Great Spice Series" was launched. Cui Yuqi, who developed as soon as possible, told me that the "fennel" used in it was inspired by the "fennel stuffed jiaozi" that I often eat at home, which is also the "taste of home" in many people’s hearts.

Last year, there was a "hand-made Schizonepeta lemon tea" in the tea, and the "Schizonepeta" used in it was also a special spice in Henan.

For regional brands, this may be a good way to build barriers. These familiar smells, on the one hand, reduce the sense of distance of new products, on the other hand, will also stimulate consumers’ curiosity.

From the brand point of view, the use of local spices can also create "collective memories" and give the brand new connotation and vitality.

2, the flavor is clear, leaving a "super memory point"

Another advantage of spices is that the memory points are clear. Spices into tea often have a clear flavor and a strong rhyme, which is easy to cause association.

For example, the round little green mint is called "toothpaste water" by consumers. This interesting name will also attract consumers’ curiosity and thus spontaneously punch in.

In the view of the person in charge of Yuanzhen’s R&D, spices have been integrated into daily life, which can make consumers have more knowledge and choose a fragrant spice with a relatively peaceful taste, which can also achieve good results.

3, style building, taking into account seasonality

Judging from the application of spices in the market, the entry of spices is now more regarded as a "style building" technique.

For example, the creation of "Southeast Asian Style" began last year, and the common gorgeous colors in Southeast Asia were added as series models by major brands. Naixue’s tea, books, and 7-cent sweet are all launched with colorful series products.

Citronella, which is often used in Southeast Asian cuisines, is also promoted by many brands, such as fragrant lemon and green lemon, and citronella has been added to bergamot lemon tea with hundreds of teas.

Cinnamon, ginger and other spices sound more suitable for "cold autumn and winter", allowing consumers to feel the warmth of winter.

In October last year, the book also introduced the Earl cinnamon tea series, and in winter, Lizhi tea also introduced a product called "Brown Sugar Ginger Tea".

Li Haoyan, director of research and development of Lizhi Tea, told me, "It’s like customers need lemon tea to get a refreshing feeling in summer, and nothing can bring a warm feeling in winter like a cup of hot ginger tea."

03 online is challenging, but it is a good idea for drainage.

In the view of many R&D, the introduction of spices into tea is still immature and faces many challenges.

Such as higher costs. Zhao Mingzhe, a senior R&D person in the industry, told me that in the production and presentation of spice drinks, syrup was used more. However, because many spices are relatively small, the production cost of factories is often high, so the price is also high, which will also increase the cost of a cup of tea.

There is also the problem of unstable flavor and quality control. "Taking citronella as an example, there are frozen raw materials at room temperature, but the reduction of citronella aroma is limited." Zhao Mingzhe said that even fresh spices have different flavors in different venues and seasons, and quality control is difficult to stabilize.

Therefore, in practical application, it is necessary to think carefully about how to compound the spice syrup with full flavor but weak freshness with fresh spices.

However, tea drinkers who are optimistic about spices entering tea are paying attention to the "marketing value" of spices. Whether it is the taste of hometown, warm feeling, or strange associations such as toothpaste water, a cup of tea has a reason for consumers to try.

In addition, some spices can also provide color. Perilla lemon jasmine tea as soon as possible, in which the application of perilla can turn tea soup into purple; And the book also burns the beautiful bodhi thick milk of the grass to upgrade the glutinous rice into beautiful glutinous rice, and the green small material brings surprises to consumers.

Color is also an important factor in selling a cup of tea. The high film production rate and the easy generation of new topics will provide consumers with new reasons to punch in.

In short, the brand value brought by using spice tea as a drainage and creative product may exceed the product itself.

tag

Finally, I want to talk about the evolution of tea-

With the development of tea industry, the meaning of products is not just "a cup of drink".

It can be a classic explosion that is sold all the year round, a niche flavor that brings surprises, a "hometown flavor" that conveys emotional links, and an innovative attempt that is somewhat risky but full of topic and discussion.

When "adding materials" makes consumers feel some aesthetic fatigue, adding spices into tea may be another feasible innovation path.

Overall planning | Laughing at every vision | Jiang Fei

　　Cctv newsAccording to the website of the Ministry of Ecology and Environment, in April, 2020, the Ministry of Ecology and Environment issued the Guiding Opinions on Implementing the Reward System for Reporting Violations of Ecological Environment, guiding all localities to establish and implement the reward system for reporting violations of ecological environment, and requiring provincial and municipal ecological environment departments to establish and implement the reward system for reporting violations of ecological environment before the end of 2020.

　　Recently, the Ministry of Ecology and Environment reported the progress and typical cases of reporting and rewarding violations of ecological environment. At present, 30 provincial-level and 313 prefecture-level ecological and environmental departments have formulated and promulgated reward regulations for reporting. In 2020, 13,870 reward cases were implemented nationwide, an increase of 44% year-on-year; The total reward was 7.19 million yuan, up 100% year-on-year. From the geographical distribution of cases, Henan, Anhui, Sichuan, Shandong and Guangdong are in the top. From the total amount of awards, Guangdong, Hebei, Henan, Anhui and Jiangsu are in the top.

　　The Ministry of Ecology and Environment pointed out that the reward system for reporting has played an important role in encouraging people to participate in the cause of ecological environmental protection, broadening the channels for finding environmental pollution problems, optimizing the law enforcement methods of ecological environment, raising the awareness of enterprises to abide by the law consciously, and promoting the realization of "three pollution control". Through the reward system for reporting, various localities have investigated and dealt with a number of major cases, which has effectively deterred all kinds of environmental violations.

　　The Ministry of Ecology and Environment said that the reward system for reporting is an important measure to optimize the law enforcement methods of ecological environmental protection and improve the efficiency of law enforcement. All localities should further promote the implementation, make full use of the reward system for reporting, further improve the pertinence and operability of this system, and use it as a bridge between the ecological environment department and the people to further enhance the people’s ecological environment satisfaction and happiness.

　　At the same time, the Ministry of Ecology and Environment also reported 10 typical cases of implementing the reward system for reporting. The eco-environment departments of Henan, Anhui, Sichuan, Shandong, Guangdong, Hebei and Jiangsu provinces, as well as the eco-environment departments (bureaus) of Tianjin, Jiangsu, Zhejiang, Henan, Guangxi and Gansu provinces (municipalities and autonomous regions), Jinjiang, Quanzhou, Fujian, Rencheng, Jining, Shandong, Dongguan, Guangdong, and Suining, Sichuan, which have achieved remarkable rewards for reporting.

　　Ten typical cases of implementing the reward system for reporting are:

　　Case 1: Tianjin Eco-environment Bureau rewarded people for reporting cases of illegal disposal of waste lead-acid batteries.

　　In July, 2020, people reported by telephone that someone illegally collected and disposed of waste lead-acid batteries at an unnamed collection point in wuqing district, Tianjin, causing environmental pollution. According to the tip-off, Tianjin Comprehensive Administrative Law Enforcement Corps, together with Tianjin Public Security Bureau, wuqing district Ecological Environment Bureau and wuqing district Public Security Bureau, responded quickly and conducted joint on-site inspection at the first time. It was found that there were dozens of tons of waste lead-acid batteries (classified as HW31 hazardous waste) at the site of the collection point. After the operators poured out some acid from the waste lead-acid batteries, they used the workshop sewer pipe to discharge them into the external environment for natural leakage. After sampling and monitoring the wastewater in the underground pipe, the pH value is less than 1, which is strongly acidic, and the total lead concentration is 10.4 mg/L. The public security organ immediately fixed the evidence and took compulsory measures against the relevant responsible persons. Tianjin wuqing district Ecological Environment Bureau investigated and dealt with the collection point according to law, and transferred its suspected environmental pollution crime to the public security organs. At present, the public security organs have arrested 44 suspects and the case is under investigation.

　　On September 27th, 2020, Tianjin Municipal Bureau of Ecology and Environment gave the whistleblower a reward of 200,000 yuan according to the relevant provisions of the Interim Measures for Reporting Environmental Violations in Tianjin and the Special Action Plan for Comprehensive Investigation and Remediation of Hazardous Wastes in Tianjin.

　　Case 2: Jiangsu Provincial Department of Ecology and Environment rewarded people for reporting a case of a factory discharging toxic substances.

　　In February 2019, the masses reported by telephone that the sewage in the electroplating workshop of an unnamed telescope parts factory in Rugao City, Jiangsu Province was directly discharged into the river. According to the clues, law enforcement officers of Rugao Ecological Environment Bureau of Nantong City conducted an on-site inspection of the factory. The inspection found that Tian rented the house of the factory to engage in the surface treatment project of the sight barrel, and discharged the untreated washing wastewater into a pond on the northeast side with a large amount of black water. The sampling and monitoring results showed that the pH value of the wastewater in the pond was 2.92, the chemical oxygen demand was 324mg/L, the total phosphorus was 46.8mg/L, and the total nickel was 1.43 mg/L. On February 25th, Rugao Ecological Environment Bureau of Nantong City took measures to seal up the factory, and transferred it to the public security organs for discharging toxic substances in the form of concealed pipes and pits, which was suspected of polluting the environment. In December 2020, Tian was sentenced to seven months in prison and fined 50,000 yuan for committing environmental pollution.

　　On May 24, 2019, the Department of Ecology and Environment of Jiangsu Province held a centralized award-winning activity for whistle-blowers of eco-environmental violations in the province. According to the relevant provisions of "Several Provisions on Protecting and Rewarding Whistleblowers of Eco-environmental Violations in Jiangsu Province (Trial)", the whistle-blower was awarded an important contribution award and was awarded 10,000 yuan.

　　Case 3: Zhejiang Provincial Department of Ecology and Environment rewarded people for reporting cases of illegal dumping of industrial solid waste across the city.

　　On May 17, 2018, the masses reported to the Zhejiang Environmental Law Enforcement Inspection Corps by telephone, reflecting that a batch of printing and dyeing sludge in Shaoxing City was transported and dumped to Jinhua City through Shaoxing Yuanteng Transportation Co., Ltd., and some license plates and parking addresses were provided. After receiving the report, the corps immediately sent law enforcement officers to Shaoxing to organize an investigation. After investigation, from 2017 to 2018, under the unified coordination of a chamber of commerce in Shaoxing, more than 30 printing and dyeing enterprises in Shaoxing entrusted the printing and dyeing sludge generated in the production process to Yiwu Lvwei Environmental Protection Technology Co., Ltd. and Lanxi Huaying Brick and Tile Factory for incineration, and entrusted a transportation company in Shaoxing for transportation. From November 2017 to June 2018, the environmental protection technology company and the brick and tile factory respectively arranged personnel to look for illegal landfill sites in the jurisdiction of Jinhua City and informed a transportation company. At night, the transportation company successively transported 26,000 tons of solid wastes containing toxic and harmful substances, such as printing and dyeing sludge, to 27 landfill sites in Jinhua City for illegal landfill, causing serious environmental pollution and direct and indirect losses of more than 50 million yuan. In October 2018, the Environmental Protection Bureau of Yuecheng District and Keqiao District of Shaoxing City imposed administrative penalties on seven enterprises involved. On May 9, 2019, the People’s Procuratorate of Yongkang City, Zhejiang Province filed a public prosecution. On November 27, 2020, the People’s Court of Yongkang City, Zhejiang Province investigated 52 individuals and 2 units for criminal responsibility for environmental pollution, and the responsible persons were sentenced to 5 years respectively. Fixed-term imprisonment ranging from one year to six months,At the same time, it was decided to bear 55.419 million yuan of social public interest loss compensation, and apologized in the newspaper.

　　The Department of Ecology and Environment of Zhejiang Province gave the whistleblower a reward of 70,000 yuan according to the relevant provisions of the Interim Measures for Reporting and Rewarding Environmental Violations of Solid Wastes in Zhejiang Province and the Implementation Rules for Reporting and Rewarding Environmental Violations of Solid Wastes in Zhejiang Province jointly issued by the former Environmental Protection Department and the Provincial Department of Finance.

　　Case 4: The Department of Ecology and Environment of Henan Province rewards people for reporting industrial wastewater cases.

　　On August 10, 2020, the masses reported to the Law Enforcement and Supervision Bureau of Ecological Environment of Henan Province that an unnamed zinc plating plant near Mingzhu Island living area in Jiyuan Demonstration Zone had illegally discharged wastewater. After receiving the report, the Law Enforcement Supervision Bureau immediately organized law enforcement officers to carry out investigations. After investigation, it was found that the zinc plating plant did not have any environmental protection procedures, nor did it build any supporting pollution prevention facilities. The zinc plating waste gas was directly discharged, and the cleaning wastewater was directly discharged into the pit on the northwest side of the plant. By sampling and testing the washing wastewater discharged from the pit and the liquid in electroplating bath, it was found that the pH value of the discharged wastewater was 3.56, the total zinc concentration was 175mg/L, and the total chromium concentration was 10.5 mg/L. The total zinc concentration and the total chromium concentration exceeded the standard by 116 times and 9.5 times respectively.

　　Jiyuan City Ecological Environment Bureau transferred the case to the public security organ for criminal responsibility. At present, the public security organ has criminally detained one person and the case is under investigation.

　　According to the relevant provisions of the Detailed Rules for the Implementation of Environmental Pollution Reporting Awards in Henan Province, the Department of Ecology and Environment of Henan Province gave the whistleblower a reward of 50,000 yuan.

　　Case 5: The Department of Ecology and Environment of Guangxi Zhuang Autonomous Region rewarded people for reporting cases of illegal dumping of hazardous wastes.

　　On the evening of January 8, 2020, the masses reported through the "12369" environmental protection hotline of the Department of Ecology and Environment of Guangxi Zhuang Autonomous Region, reflecting that Wuzhou tengxian Wuzhou Jiayuan Industrial Co., Ltd. illegally dumped and buried the waste residue produced by the factory, which seriously polluted the environment. On January 9th, the Department of Ecology and Environment of the Autonomous Region sent law enforcement officers to the scene to investigate together with Wuzhou Ecological Environment Bureau and tengxian Ecological Environment Bureau. After investigation, the company transported the generated acid-hydrolyzed black mud to a brick factory in tengxian for disposal from May to September, 2017. Due to the closure of the brick factory, a large number of acid-hydrolyzed black mud piles remained in the brick factory. In late December, 2019, the company, Yumou and Huangmou, entrusted Huangmou in heping town to treat the acid-hydrolyzed black mud in the brick factory, knowing that they were not qualified to treat hazardous wastes. From January 5th to 6th, 2020, Hwang transported the acidolysis black mud to Jigongchong Paradise Ridge in Xinliang Village, heping town for dumping and landfill, with a total weight of about 1,280 tons.

　　On February 17th, 2020, the Bureau of Ecology and Environment of tengxian, Wuzhou City transferred the case to tengxian Public Security Bureau. On December 25th, tengxian People’s Procuratorate prosecuted four suspects involved in the case. Recently, the tengxian People’s Court fined the company 1 million yuan for committing environmental pollution. Four defendants, Guimou, Adong, Ayi and Artest, were sentenced to three years’ imprisonment and fined ranging from 100,000 yuan to 50,000 yuan for committing environmental pollution.

　　On March 5, 2020, the Department of Ecology and Environment of Guangxi Zhuang Autonomous Region gave the whistleblower a reward of 10,000 yuan according to the relevant provisions of the Measures for Reporting Environmental Violations in Guangxi Zhuang Autonomous Region.

　　Case 6: Gansu Provincial Department of Ecology and Environment rewarded an environmental public welfare organization for reporting environmental violations.

　　On September 4th, 2020, an environmental protection public welfare organization reported to the Department of Ecology and Environment of Gansu Province through the Investigation Report of Jiuquan Circular Economy Industrial Park, which reflected the ecological environment, planning and water source protection of the park. Gansu Provincial Department of Ecology and Environment immediately sent a special investigation team to conduct on-site verification. The investigation team will work with Jiuquan Municipal People’s Government, Park Management Committee, planning department, water conservancy department, land department, forestry and grass department and other departments to investigate and verify relevant illegal acts.

　　After investigation, Yumen Kangbao Mining Co., Ltd. failed to build circulating pool and sedimentation tank according to the requirements of EIA, and the production wastewater was discharged into the tailings pond previously built by the enterprise; In addition, the company also has environmental violations without prior inspection. Yumen Branch of Jiuquan Eco-environment Bureau investigated and dealt with the company’s environmental violations, ordered it to correct the violations according to law, and imposed a fine of 200,000 yuan. Jiuquan Haohai Coal Chemical Co., Ltd. built a 1,000,000-ton/year tamping coke project in Yumen Dongzhen Industrial Park. The ground dust removal station did not start running, and a large amount of yellow flue gas was directly discharged during the coal loading process of No.1 coke oven. Yumen Branch of Jiuquan Ecological Environment Bureau filed an investigation on the environmental illegal behavior of the company’s unorganized flue gas emission, ordered to correct the illegal behavior according to law, and fined 180,000 yuan.

　　The Department of Ecology and Environment of Gansu Province awarded the organization a reward of 10,000 yuan according to the relevant provisions of the Measures for Reporting and Rewarding Violations of Ecological Environment in Gansu Province (Trial).

　　Case 7: Jinjiang Ecological Environment Bureau of Quanzhou City, Fujian Province rewarded WeChat for reporting the case that an enterprise discharged wastewater containing heavy metals beyond the standard.

　　On April 3, 2019, the masses reported to Jinjiang Ecological Environment Bureau of Quanzhou City, Fujian Province through WeChat, reflecting the illegal production of an unknown stainless steel door flower processing point in Qunfeng Road, Neikeng Town, Jinjiang City. After receiving the report information, Quanzhou Jinjiang Ecological Environment Bureau immediately assigned the Neikeng Environmental Protection Squadron in its jurisdiction to verify. After on-site verification, Quanzhou Jinjiang Ecological Environment Bureau set up a task force to conduct on-site investigation with Jinjiang Environmental Monitoring Station and Jinjiang Public Security Bureau Neikeng Police Station.

　　After investigation, it was found that the processing point was a stainless steel door flower processing point operated by Wen Moumou, and the environmental protection "three simultaneities" acceptance of the construction project was not handled. At the time of on-site inspection, the processing site was producing, cleaning wastewater was being discharged and there was deposition of production wastewater on the ground. Sampling and monitoring showed that the concentrations of hexavalent chromium, total chromium, copper and nickel in the production wastewater discharged from the white PVC pipeline exceeded the standard by 4-50 times respectively. The concentrations of hexavalent chromium, total chromium, copper and nickel in the ground water exceeded the standard by 31-260 times respectively. The concentration of total chromium and hexavalent chromium heavy metal pollutants in the wastewater illegally discharged from this processing point has exceeded the standard by three times, and the concentration of copper-containing heavy metal pollutants has exceeded the standard by ten times, which is suspected of environmental pollution. Quanzhou Jinjiang Ecological Environment Bureau transferred the case to Jinjiang Public Security Bureau for investigation. After the public security organ filed a case for investigation, it arrested the suspect Wen Moumou and his accomplice Zeng Moumou. In view of the fact that the discharge of wastewater containing heavy metals from the processing plant may cause damage to the surrounding environment, the public security organ entrusted a judicial appraisal institution for appraisal, and the Jinjiang Municipal People’s Procuratorate subsequently filed a criminal incidental civil environmental public interest lawsuit. At the same time, Jinjiang Ecological Environment Bureau of Quanzhou City filed a case for punishment for the illegal act of not handling the "three simultaneities" acceptance of environmental protection for the completion of the construction project, and fined 200,000 yuan.

　　Quanzhou Jinjiang Eco-environment Bureau rewarded the whistleblower with 10,000 yuan according to the relevant provisions of Jinjiang Environmental Protection Award Reporting Regulations.

　　Case 8: rencheng district Branch of Jining City Ecological Environment Bureau of Shandong Province rewarded WeChat for reporting the case of enterprises’ abnormal operation of pollution prevention and control facilities in the form of WeChat red envelopes.

　　On May 25, 2020, the masses reported to rencheng district Branch of Jining Ecological Environment Bureau of Shandong Province through the WeChat account of environmental protection prize, reflecting that Shandong Shankuang Machinery Co., Ltd. did not use the welding fume purifier in the production of welding process, and the waste gas was directly discharged.

　　After receiving the report, the person in charge of the reward for environmental protection report of rencheng district Branch of Jining Ecological Environment Bureau immediately handed over the illegal clues to the environmental law enforcement personnel of the branch. Law enforcement officers immediately went to the scene to investigate and collect evidence. On-site inspection found that the workers in the assembly workshop of the company were welding the packaging rack. Because of the large workpiece, the workers violated the management requirements when welding, and did not use the welding smoke purifier. The smell in the workshop was pungent and the exhaust gas was directly discharged.

　　Rencheng district Branch of the Bureau of Ecological Environment of Jining City, Shandong Province, according to the relevant provisions of the Interim Provisions of rencheng district on Environmental Protection Award-winning WeChat Reporting, on the day of on-site verification, 100 yuan, the whistleblower, was given a reward in the form of a WeChat red envelope.

　　Case 9: The Bureau of Ecology and Environment of Dongguan City, Guangdong Province rewarded the public for reporting a case of a company discharging toxic substances.

　　In November-December, 2019, during the trial of the reward reporting policy in Shima River Basin in Dongguan City, Guangdong Province, the masses reported that the illegal electroplating workshop was hidden in Dongguan Feitianma Plastic Hardware Products Co., Ltd., Fenggang Town, Guangdong Province, and provided the company with important clues such as locking the door at night to escape the attack, and specially setting up a back door for employees to avoid inspection. Dongguan Ecological Environment Bureau immediately contacted the public security organs and monitoring departments for careful deployment. On the evening of November 22, Dongguan Ecological Environment Bureau, together with the staff of Fenggang Public Security Bureau and Dongguan Environmental Monitoring Center Station, divided into three ways, found the entrance of illegal electroplating workshop inside the enterprise, and controlled six employees in the workshop on the spot. On-site inspection found that the workshop was equipped with 12 electroplating bath, 5 stainless steel heating tanks, and various appliances and raw materials for electroplating. The results of on-site water sampling showed that the concentrations of total chromium, total copper, total nickel, total zinc and total lead seriously exceeded the standard, which was suspected of environmental pollution crime.

　　According to the relevant provisions of the Announcement on the Trial Implementation of Rewarding Reports on Illegal Discharge of Phosphorus-containing Industrial Wastewater, the Dongguan Municipal Bureau of Ecology and Environment gave the whistleblower a reward of 500,000 yuan.

　　Case 10: Shehong Ecological Environment Bureau of Suining City, Sichuan Province rewarded WeChat for reporting the case of enterprises evading supervision and discharging pollutants.

　　On February 16, 2020, the masses reported to Shehong Ecological Environment Bureau of Suining City, Sichuan Province through WeChat, reflecting that the wastewater from Hongcheng Waste Treatment and Landfill Co., Ltd. in Shehong County was directly discharged into Fujiang River. Shehong County Environmental Monitoring and Law Enforcement Brigade immediately organized personnel to investigate. On-site inspection found that the landfill treatment company directly discharged the leachate from the collection pond into the rainwater ditch through PVC pipe with submersible pump, and finally flowed into Fujiang River. After investigation, the company discharged pollutants into the external environment in a way of evading supervision. The Shehong Ecological Environment Bureau fined the company 550,000 yuan and transferred the relevant personnel involved to the public security organs. At present, the case has been pronounced in the first instance, and the two defendants were sentenced to fixed-term imprisonment and criminal detention and fined for environmental pollution.

　　On July 1, 2020, Suining City Bureau of Ecology and Environment rewarded the whistleblower with 2,000 yuan according to the relevant provisions of the Measures for Rewarding Reports of Ecological and Environmental Violations in Suining City.