Effective school-based mental health programs are a research field with growing interest and great social value. At the stage of development and initial testing of the program, as well as during dissemination, and adaptation in other cultures, it is important that the implementation is carried out in the way that was originally intended. Fidelity or adherence is the most often used concept relating to the extent to which the implemented intervention corresponds to the originally intended program. Therefore, monitoring of the implementation is an essential element necessary to integrate into contemporary evidence-based program. The current paper describes the monitoring system developed for the Promoting Mental Health at Schools (PROMEHS) project. The monitoring was done on both the structural and procedural aspects of the program’s implementation, involving the evaluation of five core aspects: fidelity, dosage, quality, responsiveness, and adaptation. This methods article aims to describe the development of the monitoring system and to analyze the strengths of the qualitative-quantitative multi-informant approach in the monitoring of the intervention’s implementation. In the future, this would support further research on effectiveness of the PROMEHS program. Evidence connecting school-based mental health program outcomes with implementation components are increased rapidly during last years, especially in the US . It was Factors in macro-level, school-level, and individual level can affect successful program implementation in schools . SeveralIt is known that the implementation process is related to the outcomes of programs when their effectiveness is evaluated e.g., . MoreoveThere are several implementation components important for monitoring described in the literature . The criSeveral components should be included in the monitoring because we do not know which are the most important implementation factors. In previous studies, different components have been found to be the most significant implementation factors. It has been proved that interventions implemented in high fidelity show stronger effect on outcome . Recent In most cases, only few components have been assessed during the monitoring of the implementation of different preventive programs  program was developed.The Promoting Mental Health at Schools was developed within the Erasmus + Key Action 3 project co-funded by the European Commission. The project’s timeline was from 2019 to August 2022, and it aimed to develop a comprehensive mental health curriculum, implement it, and evaluate its effectiveness. The consortium involved researchers, practitioners, and policymakers from seven European countries: Italy, Latvia, Portugal, Croatia, Romania, Greece, and Malta.The PROMEHS theoretical framework includes three domains, namely, promoting social-emotional learning (SEL) and resilience and preventing social, emotional, and behavioral problems. This framework was described and substantiated by The key features of the universal curriculum were based on principles of international research , such asThe PROMEHS curriculum consists of seven handbooks. Two are for teachers with ready-to-use, step-by-step activity plans for leading pre-school and school students aged from 3 to 18 years. Two handbooks are for both pre-school/primary school students and middle/secondary school students with activities to carry out independently at home or together with their parents. The other three handbooks are for teachers to promote their own mental health, for parents to promote mental health at home, and for supplying recommendations to policymakers.Since the curriculum was aimed at fostering students’ SEL and resilience and preventing social, emotional, and behavioral problems, all these topics were covered in the offered activities. Each activity has the same structure, namely, defined learning outcomes, a clearly defined age group, and a step-by-step activity plan. The activity starts with a story, followed by a discussion, role-play, group work, or another learning strategy. An important part of the activities is reflection. At the end of every activity, a teacher is provided with a brief formative evaluation chart, tips on how to embed the goal into their everyday teaching practices, as well as culturally adapted further resources .The curriculum was implemented in Italy, Latvia, Portugal, Croatia, Romania, and Greece, whereas the University of Malta acted as the external evaluator and was not involved in the development and implementation of PROMEHS. The project’s implementation and the evaluation of its effectiveness were carried out in four age groups of students from pre-school to secondary school level , including disadvantaged children.The quasi-experimental research design with experimental and control conditions was implemented to evaluate the effectiveness of the program. An integral part of the development and implementation of the PROMEHS program was the monitoring system, which was built with the purposes of ensuring the fidelity and quality of its implementation and of finding out culture-specific practices to develop recommendations for practitioners and educational policymakers see . A detaiThe monitoring of the implementation can be done for diverse purposes, and decisions should always consider the balance between costs and added value. In this case, the purpose of the monitoring, as defined in the project proposal, was to evaluate the quality of the intervention’s implementation (1) to ensure the fidelity and quality of its implementation and (2) to find out culture-specific practices for schools to develop recommendations for both practitioners and educational policymakers.Five dimensions were used for this purpose: fidelity, dosage, quality, responsiveness, and adaptation .Fidelity characterizes the extent to which the implemented intervention corresponds to the originally intended program. The fidelity of the implementation of the PROMEHS program was supported by the provision of detailed materials on the content and procedures to be implemented. Both structural (the content to be delivered) and process components (how the content should be implemented) of the program’s implementation were described in the PROMEHS materials. Comprehensive and detailed handbooks were developed for each age group, both for teachers and students ,b,c,d, tDosage refers to how much of the intervention has been delivered. It has a high potential to be included in effectiveness studies, and therefore it was decided to monitor it as well. In the implementation of the PROMEHS, minimal exposure was defined as 12 activities proportionally covering all three parts of the program, namely SEL, promoting resilience, and preventing behavioral problems.Quality refers to how well different program components have been implemented. The quality of implementation evaluates the competence of the program providers according to the content and manner of the intervention manual. Organizational factors  are recognized as a useful way to increase the quality of the intervention. However, its combination with process evaluation is crucial, especially when using external observations .Participant responsiveness refers to the degree to which the program stimulates the interest and engagement of participants. Most often, it is the responsiveness of the direct target group  that is measured . ConsideConsidering that the model for assessing the fidelity of the PROMEHS project’s implementation was developed to provide information on how its implementation may vary across countries and to provide specific recommendations for its implementation in the future, fidelity is supplemented by adaptation assessments. Adaptation refers to changes made to the original program during its implementation (program modification). Previous research  suggestsThe PROMEHS program was implemented in six European countries in the school year 2020/2021. Initially, it involved 10,209 students, but pre- and post-test evaluations were received from 4,501 participants in the experimental condition and 3,288 participants in the control group, where the evaluators were teachers. Both pre- and post-test parental evaluations were received in relation to the outcomes of 2,394 participants in the experimental group and 2,234 participants in the control group. Student self-reports at the two measure points were obtained from 1,845 students in the experimental group and 1,458 from the control group.However, monitoring the data collection was not directly related to pre- and post-test data for the effectiveness study. The monitoring sample consisted of experimental condition participants, namely 2,534 students from primary and middle/secondary school (aged nine and older) and 2,868 parents, who provided feedback after their children’s participation in the program activities See .During the project’s implementation, 532 teachers were trained in total, of whom 421 filled out the final evaluation of the program, identifying the strengths and weaknesses of the materials and providing practitioners with their expertise for the further elaboration of the PROMEHS materials.School support teams were organized in each country, with a range of members from three to eight per country. In sum, there were 29 members, all qualified professionals with specific knowledge and expertise as described in the quality requirements. They organized pre- and post-test data collection, managed teacher training and on-going supervisions, collected qualitative data from teachers, and contributed to developing detailed recommendations for the further elaboration of the PROMEHS program’s materials and its implementation in diverse contexts .The number of participants was different between the countries due to different response rate (in groups of students and their parents), and the teachers’ involvement .Several steps were taken for the development of the monitoring system as recommended in the literature : (1) defThe monitoring system was developed by the first two authors of the paper in collaboration with project partners. Considering the purpose of the monitoring and principles of the program, a multi-component and multi-informant approach was chosen.Detailed implementation procedures were developed following recommendations in literature . EssentiFollowing examples in the previous studies e.g.,  and guidAll procedures characterizing process components (how the program should be implemented) were discussed with project partners and translated into six national languages. All measures were piloted with the appropriate target audience, tested and corrected for clarity, discussed with partners, and translated into six national languages. Finally, monitoring data were collected during all stages of the field trial and analyzed before the results were presented to project partners.The Ethics Committee for Humanities and Social Sciences Research Involving Human Participants of the University of Latvia granted permission for the research on 12 December 2019.The PROMEHS monitoring system resulting from the procedures and measures developed for monitoring purposes can be seen in The PROMEHS monitoring procedures are presented in The essential components of the monitoring were selected following curriculum and research protocols and were described in the framework section. Materials and guidelines were available for the school support team, sharing information on data collection and training, meeting with school leaders, teacher training and supervisions, activities at schools, and parents’ meetings.Aiming to evaluate the fidelity of the program’s implementation, data were collected after teacher training, during activities at school, and during supervisions from both school support team members and teachers. Measures included categorical scales  as well as continuous scales with a Likert-type scale , provides the opportunity to test the dosage effect in relation to the effectiveness of the PROMEHS program.The quality of implementation evaluates the skill and competence of the program providers according to the content and methods of the PROMEHS intervention manual. There were specific competence requirements for school support team members, and regular meetings related to testing, training, and supervisions were organized and reported. Several support materials were provided to strengthen their competence in PROMEHS materials  and manaThe quality of evaluation addresses school support team members (self-reports) and teachers (self-reports and student reports). School support team members evaluated their own competence in teaching materials and procedures  before sConsidering the principles of the PROMEHS program emphasizing collaboration between school and family, teachers, students, and parents were all treated as participants, and their levels of responsiveness were measured. Teacher responsiveness was estimated after the teacher training and was evaluated by school support team members . StudentAdaptation refers to changes made to the original program during its implementation . As a result, adaptation was integrated as an independent dimension with high value in the monitoring system in all stages of implementation, and qualitative data were collected. Adaptations were observed in several sources. Teachers filled out a self-reflection form  after eaA program can be evaluated as effective if it is implemented as intended. The fidelity of the intervention can be significantly increased by developing materials on content, what to implement, and the manner in which it should be implemented. The PROMEHS program filled this requirement by providing comprehensive, ready-to-use handbooks for teachers, students, families, and policymakers. Added value is related to the inclusion of content and process components in the monitoring system, where several materials can be used as tools to familiarize oneself with the content of the program while following the guidelines described in detailed procedures. Additionally, the monitoring measures include a checklist to ensure the implementation of all program components.The PROMEHS monitoring system covers all the most relevant components of the intervention, starting with the development of the school support team, followed by teacher training, supervision, activities at schools, parents’ meetings, and student feedback. Specific requirements for the school support team members are described in the monitoring procedures to ensure quality. Moreover, these prepared the professional continuing education course for pedagogues so that PROMEHS could be maintained sustainably.Providing support for people involved in the implementation of the program was recognized as a crucial principle, and therefore materials in the form of checklists were included to make the monitoring system user-friendly and helpful, allowing participants to practice self-monitoring during the implementation. A checklist on the content of handbooks allowed implementers to use it both as a training task and as a piece of evidence on how familiar both the school support team and teachers were with the provided materials. Considering that the usefulness and user-friendliness of tools can increase readiness to use monitoring tools, further research is needed on the applicability of the monitoring system after the project. It can be assumed that teacher self-report forms e.g.,  can be uThe implementation of the program is not always compatible with an aim to explore factors affecting its success or failure. A research strategy combining monitoring data and pre- post-test data allows the testing of a hypothesis about possible mediating or moderating effects of implementation characteristics on program outcomes. It can be assumed that diverse informants can evaluate different qualities of the program’s implementation, allowing key predictors of program efficacy and necessary support for program providers to be explored.Both quantitative and qualitative , since the intervention included activities focused on teachers, parents, and school-leaders as well. Direct observation was not included in the present monitoring system; however, this limitation was addressed by collecting multi-informant data from the program’s implementers (teachers), students, and their parents, as well as from the support team members, who provided teacher training and on-going supervisions and parents’ meetings. This strategy allows the implementation of the community engagement principle to be monitored as well, which is crucial to the sustainability of the program.The findings of this study highlight the importance of including several aspects often generally described as fidelity but which, nevertheless, allow the implementation process of a program to be evaluated from different angles, namely, dosage, responsiveness, quality, and adaptation.This study also emphasizes the role of monitoring every aspect of implementation regarding both its content and its procedure. Moreover, it emphasizes the importance of building scientifically sound and, at the same time, user-friendly monitoring procedures in order not to overwhelm participants with data collecting but rather to support them during the implementation process. This study contributes to the field providing an elaborated framework for monitoring of implementation of different interventions. This supports both researchers and practitioners in developing, implementing, assessing, and sustaining the best possible practice in the intervention.The strengths of this monitoring system are its observation of both content and process with scientifically sound dimensions, thus covering the whole spectrum of implementation, its collection of qualitative and quantitative data, and its use of a multi-informant approach. PROMEHS implementation during the COVID-19 pandemic allowed to document adaptation related with remote learning and computer mediated instructions.The system also has some limitations. First, no direct observation of the teacher’s competence and interaction with students during the activities was available, limiting conclusions about the quality of the implementation of the program. Observation would be beneficial for providing more contextualized feedback and helping to develop teacher competence in instructing SEL. However, this can partly be offset with observations during supervisions when teachers interact with each other, which can also be used as an indicator of the manner in which they implement principles of the PROMEHS program. This limitation was partially neutralized by collecting evaluations from all groups of participants, direct observation of the responsiveness during teacher training was done by school support teams, whereas responsiveness of the students was evaluated by their parents. In the future, it would be useful to add direct observation during class activities to estimate quality of the implementation, as well as responsiveness of the students. Second, there was no monitoring of the control group. One critical point that was emphasized in the literature was the necessity to control other possible interventions in the control group. However, the COVID-19 pandemic context, with the related social distancing and remote learning, provided an opportunity to overcome this limitation since, due to the restrictions of the pandemic, the control group did not receive any alternative interventions. This naturally alleviated the necessity to monitor it.The original contributions presented in this study are included in the article/The studies involving human participants were reviewed and approved by the Ethics Committee for Humanities and Social Sciences Research Involving Human Participants of the University of Latvia on 12 December 2019. Written informed consent to participate in this study was provided by the participants or their legal guardian/next of kin.BM: lead writer, arranging the research in Latvia, contributing to the development of the monitoring system, and collecting data. IS: contributing to the development of the monitoring system and to writing. IG: a key contribution to designing the research and revising the manuscript. All authors contributed to the article and approved the submitted version.


Mechanism study revealed that CD229 promoted MM cell proliferation by regulating the RAS/ERK signaling pathway. Further exploration employed co-immunoprecipitation coupled with mass spectrometry to identify RASAL3 as an important downstream protein of CD229. Additionally, we developed a co-culture method combined with the immunofluorescence assay to confirm that intercellular tyrosine phosphorylation mediated self-activation of CD229 to activate RAS/ERK signaling pathway via interacting with RASAL3. Taken together, these findings not only demonstrate the oncogenic role of CD229 in MM cell proliferation, but also illustrate the potential of CD229 as a promising therapeutic target for MM treatment.Multiple myeloma (MM) is an incurable plasma cell malignancy, while CAR-T therapy offers a new direction for the treatment of MM. Recently, signaling lymphocytic activation molecule family 3 (CD229), a cell surface immune receptor belonging to the signaling lymphocyte activating molecule family (SLAMF), is emerging as a CAR-T therapeutic target in MM. However, a clear role of CD229 in MM remains elusive. In this study, MM patients with elevated CD229 expression achieved poor prognosis by analyzing MM clinical databases. In addition, CD229 promoted MM cell proliferation  Multiple myeloma (MM) is a hematological B-lymphocyte malignancy with clonal expansion of plasma cells in the patient's bone marrow, which can ultimately cause hypercalcemia, renal failure, anemia, bone disease and immunosuppression , 2. MM pIt is of note that chimeric antigen receptor T (CAR-T) cell therapy has been already introduced for treating MM in clinical trials –11. ReceInspired by the current studies on the effective efficacy of CD229-based CAR-T therapy, we herein examined the expression of CD229 in a subpopulation of myeloma, and demonstrated the role of CD229 in promoting MM cell proliferation through the underlying mechanism of CD229-mediated RAS/ERK signaling. It is suggested that CD229 may be served as a novel biological target for treatment of MM.p < 0.001) (p < 0.05) (p < 0.05) (p < 0.05) (p < 0.01) (p < 0.01) , monoclonal gammopathy of undetermined significance  and MM patients  (gene expression dataset GSE5900). The results showed that the CD229 mRNA level was significantly increased from NP, MGUS to MM patients during disease progression (< 0.001) . Intrigu < 0.05) . In deta < 0.05) . High ex < 0.05) . Further < 0.01)  and rela < 0.01) . Collectp < 0.01) (p < 0.01) (p < 0.001) (p < 0.01)  in MM cells through lentiviral transfection, as confirmed by Western blotting (WB) , 1B. MTT < 0.01) . Cell cy < 0.01) . Inverse < 0.01) . CD229-K< 0.001) . In addi < 0.01) . Taken tTo further explore the mechanism by which CD229 affected MM cell proliferation, we prepared RNA samples for transcriptomic RNA sequencing (RNA-seq) to screen differentially expressed genes between CD229-OE cells and their corresponding WT cells. In total, we found that 236 genes were upregulated and 254 genes were downregulated in CD229-OE MM cells compared to WT cells . As showin vivo, the xenograft mouse model was constructed in the immunodeficient NOD/SCID mice by subcutaneous injection of WT and CD229-OE MM cells into the left and right flank of mice, respectively. Interestingly, we observed that the volume of the abdominal tumor formed by CD229-OE MM cells appeared larger than those formed by WT MM cells (p < 0.01)  . To furtWT group , suggestTo further explore how CD229 activated RAS, we performed Co-IP coupled with MS (Co-IP/MS) to screen the client proteins. Among these CD229-regulated proteins see , 5B rankIt has been reported that CD229 signal transduction requires phosphorylation of the immune receptor tyrosine signal motif (ITSM) , 25. We high T cells during activation while spare functional CD229neg/low T cells without abolishing the cytotoxicity of T cells [MM remains a life-threatening and incurable plasma cell malignancy, while CAR-T therapy has emerged as an effective method and the curative effect from CAR-T clinical trials demonstrate a better efficacy with impressive overall response rates. Rational utilizing of targets is a prerequisite of CAR-T cell therapy. Recently, the cell surface receptor CD229 is developed as a novel and alternative CAR-T therapeutic target for MM. While the presence of CD229 on both B and T lymphocytes occurred, the CAR-T cells targeting CD229 were highly active against memory B cells and MM-propagating cells but exhibited no fratricide during CD229 CAR-T cell production. It may partially due to that the protein expression of CD229 is down-regulated in activated T cells compared to normal T cells, the CD229 CAR-T cells target normal CD229 T cells , 28. In  T cells . In presin vitro and in vivo xenograft mouse model. However, elevated CD229 expression inhibited cell proliferation, migration and induced apoptosis in hepatocellular carcinoma [First, we verified that the elevated expression of CD229 was linked to poor prognosis in MM patients, especially in the highest risk PR subgroup with the highest degree of malignant proliferation. Our study validated that CD229 expression was significantly associated with the malignant proliferation of MM cell lines arcinoma . It is pin vivo tumor tissues, supporting their universal expression and critical regulatory role in cellular proliferation. Furthermore, RASAL3 was screened and validated as an intermediate protein between CD229 and RAS through Co-IP/MS experiment. As the most recently identified Ras GTPase activating protein, RASAL3 can accelerate RAS-GTP hydrolysis to form RAS-GDP, thereby functioning as a brake on RAS signaling pathway [In present study, the specific mechanism of CD229 in promoting MM cell proliferation was screened and validated by transcriptome sequencing and Co-IP/MS analysis. Among the top-ranked pathways, the enrichment correlation of endocytosis and the lysosome signaling pathway might reflect the role of CD229 as a membrane protein. Furthermore, the primary immunodeficiency signaling pathway was in accordance with the participation of CD229 in the systemic lupus erythematosus (SLE), an autoimmune disease . More im pathway . These rCD229, as a cell surface receptor, is activated by receptor-ligand interaction , 40. It in vitro and in vivo. Mechanistically, CD229 interacts with RASAL3 upon tyrosine phosphorylation-mediated self-activation, thus activating the RAS/ERK signaling pathway. Our study not only demonstrates the oncogenic role of CD229 in MM cell proliferation, but also illustrates the new theoretical basis on CD229 as a promising therapeutic target for the treatment of MM.In conclusion, the present study reveals that elevated CD229 expression confers poor survival in MM patients and promotes cell proliferation both CD229 mRNA was determined using the gene expression profiling (GEP) cohorts, which were mined from the GEO database as previously described , 48. TheThe primary antibodies used in this study were at the dilutions of 1:1000 as follows: CD229 ; RASAL3 ; Flag ; ERK ; p-ERK ; GAPDH . The second antibodies Goat anti-Rabbit IgG (H+L) HRP  and Goat anti-Mouse IgG (H+L) HRP  were in 5000 diluted concentrations. Puromycin was obtained from Merck KGaA . Diphenyltetrazolium Bromide (MTT) was purchased from Solarbio . PKH26 was purchased from Sigma-Aldrich .2 incubator.Human MM cell lines ARP1, CAG and peripheral blood mononuclear cells (PBMCs) were cultured in RPMI-1640 . HEK293 cells were cultured in DMEM . Culture medium was added with fetal bovine serum , penicillin  and streptomycin , which was changed every 2 days. All cells were cultured in 100 mm dishes at 37° C in the 5% COPlasmids containing human CD229 cDNA were provided by TranSheepBio  and CD229 siRNA were synthesized by GenePharma . The CD229 cDNA was cloned into the lentiviral vector, CD513B-1, and linked with FLAG tags. Lentiviruses containing cDNA were obtained by co-transfection of CD229 expression vector with packaging vectors  using Liposomal Transfection Reagent  . The virThe cells were resuspended with electroporation solution. Subsequently, siRNA was added into the solution to a final concentration of 100 nmol/L and then the solution was transferred into the electroporation cuvettes plus. Two pulses for 1.0 s at 960 microF of capacitance, 200 V of voltage were the most favorable electrical parameters for efficiency.Cells were cultured in 96-well plates at a density of 2,000 cells/well. The Thiazolyl Blue Tetrazolium Bromide (MTT) method was performed to test the proliferation rate and cell viability for 24, 48 and 72 h, respectively. Absorbance was read at 570 nm using the microplate reader.The cell cycle was detected by flow cytometry  as previously described .Protein levels were determined by WB analysis under the protocol as previously described (50). According to the manufacturer’s instructions, the Pierce Direct Magnetic IP/Co-IP kit  was utilized for Co-IP assays. As CD229 cDNA is linked with FLAG tag, the FLAG antibody was used instead of the CD229 antibody for IP. And the IgG antibody sharing the same immunogen with the IP antibody was chosen as a negative control.RAS activity was detected according to the instructions of the Active Ras Pull-Down and Detection Kit .6 CAG CD229-OE cells were injected subcutaneously into the left abdomen of mice and the same amount of CAG WT cells were subcutaneously injected into the right abdominal cavity of mice. The diameter of the tumor was measured using a vernier caliper every 1~2 days. When the xenograft tumor diameter was up to 15 mm, the mice were sacrificed, and then the tumors were collected, weighed and photographed.MM xenograft model was established in 6~8-week-old SCID/NOD mice. Briefly, 1 × 10SDS-PAGE was used to separate proteins from CD229-OE cells. Gel bands at the expected size were excised and digested with sequence-grade trypsin . The proteins were first quantified, and followed by reductive alkylation to open the three-dimensional structure of the proteins. The peptides were extracted by enzymatic digestion and analyzed by MS . Finally, the peptides were analyzed according to National Center for Biotechnology Information nonredundant protein database .Immunofluorescence staining experiments were performed as previously described . Images p < 0.05 was considered statistically significant.All data were expressed as the mean ± standard deviation. Two-tailed Student’s t-test (2 groups) or one-way ANOVA for multiple comparisons were used to determine the significance between experimental groups. The Kaplan–Meier method and Log-rank test were used to determine the survival rate of MM patients. *All supporting data are included in the manuscript and available upon reasonable request to the corresponding author. The original contributions presented in the study are publicly available. This data can be found here: the ProteomeXchange Consortium: PXD032177. The RNA-seq data was deposited in GEO (GSE199200).Supplementary Figure 1Supplementary Table 1Supplementary Table 2


Integr Environ Assess Manag 2022;18:407‐427. © 2021 The Authors. Integrated Environmental Assessment and Management published by Wiley Periodicals LLC on behalf of Society of Environmental Toxicology & Chemistry (SETAC).Historically, environmental research and monitoring in the Alberta oil sands region (OSR) located in northeastern Alberta, Canada, have largely neglected, meaningful Indigenous participation. Through years of experience on the land, Indigenous knowledge (IK) holders recognize change on the landscape, drawing on inextricable links between environmental health and practicing traditional rights. The cumulative impacts of crude oil production are of great concern to Indigenous communities, and monitoring initiatives in the OSR provide unique opportunities to develop Indigenous community‐based monitoring (ICBM). A review of ICBM literature on the OSR from 2009 to 2020 was completed. Based on this review, we identify best practices in ICBM and propose governance structures and a framework to support meaningful integration of ICBM into regulatory environmental monitoring. Because it involves multimedia monitoring and produces data and insights that integrate many aspects of the environment, ICBM is important for natural science research. ICBM can enhance the relevance of environmental monitoring by examining relationships between physical and chemical stressors and culturally relevant indicators, so improving predictions of long‐term changes in the environment. Unfortunately, many Indigenous communities distrust researchers owing to previous experiences of exploitive use of IK. In the present paper, we recommend important practices for the integration of IK into regional environmental monitoring programs. ICBM is important to communities because it includes conditions to which communities can exercise traditional rights, and highlight how industrial activities affect this ability. Equally important, ICBM can generate a resurgence of Indigenous languages and subsequently traditional practices; it can also revive the connection with traditional lands and improve food security.  The paper provides a unique way of organizing the integration of Indigenous knowledge (IK) and Western science (WS) using a conceptual model, which can allow regional monitoring programs to constantly evaluate research objectives and identify gaps.It is not the intention of this paper to adapt IK or WS to any model or monitoring program, but to discuss the integrative ability of both to better understand pressure/stressor‐pathways and response relationships of environmental impacts in the oil sands region.Contextualizing each program into the conceptual model is useful for large monitoring programs, such as the Oil Sands Monitoring (OSM) program, to integrate large amounts of data assessing different aspects of the environment over time and space.Indigenous community‐based monitoring approaches, such as those led by the Mikisew Cree and Athabasca Chipewyan First Nations, support regional monitoring where data are easily captured in regional‐scale models. The oil sands region (OSR) located in Northern Alberta, Canada, encompasses approximately 11 million acres (4.3 million hectares), an area roughly the size of the US state of Florida. The ground beneath this region is a mixture of sand, clay, and a heavy crude oil or tarry substance known as bitumen, collectively referred to as tar sands or oil sands. Considerable amounts of water and land are required for oil sands extraction. The consequences to the Athabasca River watershed have been the focus of several decades of environmental monitoring by the provincial and federal Canadian governments, as well as industry. Industrial development creates concerns among Indigenous peoples regarding the safety of consuming traditional resources, such as wild game, and the quality of water and air . ICBM utilizes both Indigenous knowledge (IK) and Western science (WS), combining the strengths of both to tackle challenging environmental issues. Lessons can be learned from experiences involving Indigenous people in the planning, development, and integration of ICBM into regional monitoring programs. In the OSR and elsewhere in Canada, IK is maintained, transmitted, and developed by Indigenous peoples through lived experience on the land, exercising Section 35 Rights of the Constitution Act of Canada  over many generations  released 94 Calls to Action to help facilitate reconciliation between Indigenous and non‐Indigenous peoples in Canada TRCC, . AlthougIn 2017, the governments of Alberta and Canada included a commitment to enhance the role of ICBM in the oil sands regional monitoring program with improved capacity building and a memorandum of understanding . Working collaboratively with the governments of Alberta and Canada, 18 Indigenous communities (and regions) in the OSR subsequently participated in the development of an Operational Framework Agreement (OFA). The OFA, developed to guide the governance of the joint Alberta–Canada Oil Sands Monitoring (OSM) program, strives to improve Indigenous participation, transparency, and inclusion of IK  to help facilitate Indigenous participation in OSM programs and advise a Science and Indigenous Knowledge Integration Committee (SIKIC). The ICBMAC can play a key role in fostering ethical integration of IK and community‐developed, culturally relevant indicators in OSM programs, including sociocultural and sociopolitical components that typically fall outside WS monitoring programs, such as accessibility of Section 35 Rights. The SIKIC, in turn, evaluates recommendations for funding OSM programs, including ICBM and capacity‐building programs, and provides annual recommendations to the OSM program cochairs on funding allocation . Further, to ensure consistency and the ability to integrate data across a regional monitoring program, capacity can also refer to the ability to follow specific standardized quality assurance and quality control (QA/QC) and standard operating procedures (SOPs). However, capacity in Indigenous program delivery can and should also be defined as experience with ICBM methodologies, local knowledge, scientific skills, safety skills, technical skills, communication skills, holistic oversight, and project management , they can still offer a starting point to increasing capacity in communities who may want to develop their own ICBM programs that use both IK and WS. Best practices in ICBM will encourage meaningful opportunities for community members, especially youth, to participate in science, as this can facilitate capacity for ICBM over generations  and the Athabasca Chipewyan First Nation (ACFN). The MCFN and ACFN live downstream of oil sands industries, along the Athabasca River and into the Peace–Athabasca Delta (PAD). As a complex and ecologically diverse freshwater delta, PAD is very important to Indigenous communities. The MCFN, discussed next, are well known for their involvement in environmental monitoring strategies in the OSR and have greatly influenced ICBM and the integration of IK in regional monitoring practices  for approval of the Frontier Oil Sands Project. The project request included construction, operation, and reclamation of an existing oil sands surface mine located approximately 30 km south of Wood Buffalo National Park , specific to the region, as potentially the last remaining Wood bison unaffected by disease spread through the introduction, many years earlier, of Plains Bison (Bison bison bison) by authorities . The severity of environmental concerns detailed in the MCFN petition received enough support from other Indigenous communities, environmental organizations, and scientists to attract attention to the issue . Specifically, the MCFN identified a unique herd of bison, the Ronald Lake bison  undertake a strategic environmental assessment (SEA) to assess the potential cumulative impacts of various developments, including oil sands, on Wood Buffalo Nation Park (World Heritage Site) and PAD. The completion of the SEA was a collaborative effort between governments, researchers, nongovernmental environmental organizations, and Indigenous communities in and around the Wood Buffalo National Park . The SEA involved extensive information gathering including IK and WS, intended to inform project‐level environmental assessments and to influence the development of an action plan to protect Wood Buffalo National Park . The letter draws on key principles that extend beyond environmental impact assessments, including perceived quality and quantity of resources, as well as land accessibility to know and teach Indigenous law and practices of environmental stewardship . At that time, the MCFN also presented a letter to the Chair of Frontier Oil Sands Mine, identifying the necessity of assessing impacts on the exercise of Aboriginal and Treaty Rights .The principles raised by the MCFN for the Frontier project, along with the 10 Calls to Action proposed by Wong et al. ; Table 1Conceptual models are designed to be tools for organizing and communicating knowledge, and can offer space for mutual understanding among holders of traditional, local, and scientific knowledge Suter, . This ICOverall, the scope of OSM programs is limited to impacts related to oil sand, which must be linked to a conceptual model and organized along a pressure/stressor‐pathway and response continuum  landscape disturbances ; and (2) contamination . Changes to the land influence the environment through multiple stressors and pathways, resulting in multiple idiosyncratic effects and responses, terrestrially and aquatically, that continue to challenge mitigation efforts . Acute air quality refers to the quality of air examined that may be harmful when exposure durations are minutes to hours. The conceptual model  in response to concerns raised by the Fort McKay First Nation (FMFN) related to oil sands development  events, which can result in dangerous short‐term air quality, odors, and haze within the community. The AER received 172 air‐quality complaints from Fort McKay community members between January 2010 and December of 2014. Owing to complaints, aligning emergency response air‐quality exceedances and odor complaints became a priority to the AER  was launched in February 2013, with the objective of understanding the relationships between odors experienced in the communities and oil sands development Dann, . To ensuIn the first year of the program, more than 150 odor days (days with seven or more air odor complaints) were reported, growing to more than 270 the following year, peaking both years in summer WBEA, , 2015a. Although not classified as ICBM, opportunities to participate in WS projects such as COMP can provide opportunities for Indigenous communities to participate in monitoring activities and address concerns related to air quality. These experiences can build capacity within the community to develop Indigenous‐led programs involving air quality and odors. Increasing community involvement in regional air‐quality programs over time can increase the amount of air‐quality data in the OSR and in remote locations that may only be easily accessed by Indigenous community members who live there. Ensuring the credibility of multiple datasets will take careful attention and training in WS practices where applicable . WS programs involving Indigenous peoples can also be integrated with other ICBM programs in the OSM program. Improving our understanding of stressor/pressure‐pathway‐response relationships in the OSR will involve linking air‐quality data (pressure and stressors) with responses in the environment, such as the impact of contaminant deposition on food quality and the perception of food safety. For example, the Indigenous‐led Berry Harvesting project, part of WBEA's Integrated Atmospheric Pollutant Deposition Monitoring Network, discussed next, focuses on dry deposition and specific atmospheric pollutants that could endanger the environment and culturally significant resources WBEA, .WBEA's Integrated Atmospheric Pollutant Deposition Monitoring Network is designed to detect and quantify atmospheric pollutant deposition to the terrestrial environment, which includes impacts on traditional land and resources WBEA, . As showIndigenous communities living in the OSR have their own culturally relevant way of observing the effects (response) of emissions on the land because they have long‐standing IK that allows knowledge holders to recognize change  in Wabasca Lake. Community members were worried about safety of fish consumption owing to abnormalities observed in the fish, which included visible tumor‐like abrasions on fish skin. The goals of the project were to address gaps in water‐quality knowledge and information about North Wabasca Lake and to provide monitoring training in scientific sampling methodology to Bigstone Cree Nation  during summer (June–September) and winter (February). Ten members from Bigstone Cree Nation participated in safety and sample collection workshops. Overall, the work revealed no metal contamination derived from human activity in the watershed. In addition, skin tumors on recently caught walleye that concerned community members were determined to be caused by a natural virus known as Lymphocystis, commonly found in Alberta's lakes. The virus is not usually fatal to fish and is not known to affect humans and other mammals . PACs represent a broad range of chemicals, including polycyclic aromatic hydrocarbons (PAHs). PAHs are organic hydrophobic compounds released from many sources in the Lower Athabasca River system, some naturally from oil sands deposits and others from extraction and upgrading facilities, forest fires, urban runoff, and industrial activities. Consequently, PAHs can be taken up by plants, accumulate in sediments, and can be found in tissues of terrestrial and aquatic organisms  and ecologically complex freshwater deltas in the world, filled with rivers, lakes, channels, marshes, and grasslands  established guidelines to identify national water‐quality criteria for drinking water, communities may not agree with these thresholds CCME, . CurrentOverall, as a community‐led ICBM program, community members can be empowered to answer specific questions that are meaningful and important to them. Increasing capacity within communities to develop ICBM to monitor the quality of water can also provide security and access to traditional resources. ICBM can also improve transparency of data results and potentially contribute to OSM's regional surface‐water‐quality data repositories, including sampling in remote locations that may not be easily accessed by non‐community members. Currently, the MCFN place their water‐quality data on the Makenzie Data Stream (2021) database, which is openly accessible. The OSM Surface Water program currently uses KISTERS (2021) to house OSM data; however, it may not be recognized by communities or support community needs to store sensitive information. In the past, issues involving the collection, storage, accessibility, and usage of ICBM data, especially if the data contains sensitive information such as sacred IK, have been an issue for communities participating in regional programs  is braided with WS , can help improve understanding of whitefish health and provide communities the ability to monitor culturally significant resources. Braiding IK and WS regarding the health of fish can address the OSM program's objective to document change and potentially begin to link change to specific development activities, such as oil sands. Using the conceptual model Figure , changesIndigenous communities in the northeastern oil sands region, in particular the Fort McMurray Métis, are concerned about the disappearance of freshwater clams in the Athabasca River. IK holders in the region reported that freshwater clams have declined and, in some areas of the Lower Athabasca, have completely disappeared over the past 20–40 years. The Fort McMurray Métis rely on and are intimately connected to these waterways for navigational purposes and for traditional resources, which for generations has included harvesting freshwater clams  can disproportionately influence navigability than if the river had higher discharge volumes  from traditional territories and traplines. Tissue samples were sent for lab analyses to assess contaminant burdens. Semi‐aquatic species were chosen because they are closely related to water and are good indicators of ecosystem health. With a diet consisting predominately of fish, semi‐aquatic wildlife may be exposed to contaminants in the water and might accumulate toxins in muscle and organ tissues, especially the liver. The selection of semi‐aquatic species was made by scientists; however, which species, where, and when was guided by community members. The lab analyses of collected samples from participants indicated that wildlife collected closer to Fort McMurray industrial activities had higher levels of contaminants than wildlife collected farther away  and Wood Bison (Bison bison athabasca). The cultural importance of wetland plants can include many things such as food resources, medicinal products, and support for spiritual values. However, the IK acquired to discover these uses is a result of accessible, abundant, and biologically diverse wetland ecosystems over generations  atmospheric deposition (pathway) and the development of indicators to detect deposition from oil sands emissions (responses), (2) effects of hydrological alteration (pathway) and the development of indicators to detect alteration patterns (responses), and (3) impacts of land disturbance (stressor) and the development of indicators to detect changes on wetlands (responses). Each area involves the possibility for integration with other OSM programs focusing specifically on those theme areas. The primary goal of this program is to develop a long‐term wetland monitoring program that includes and applies IK related to wetlands and ecosystem change. As part of this goal, the program is integrated with the Culturally Important Wetlands Plants program. The objective of the Culturally Important Wetlands Plants program is to provide space and resources for communities in the OSR to share IK and develop ICBM programs that are culturally meaningful . It remains to be seen how the OSM program will move these objectives forwards; however, the development of the programs is a good indication the OSM program is heading towards an integrated strategy that combines the strengths of IK and WS to tackle challenging environmental issues.After the Culturally Important Wetlands Plants program moves past the pilot phases, monitoring health and abundance of indicator  species in wetlands can improve understanding of impacts associated with contaminants, landscape disturbance, or a combination of both on wetland ecosystems, which ultimately challenge the meaningful exercise of Section 35 Rights  can alter behavior of Indigenous peoples because of concerns over the access and safety of traditional resources from these environments. This could prevent some community members from exercising their Section 35 Rights and passing IK on to the next generations. To participate, collaborate, or develop ICBM programs, Indigenous participants must be able to practice Section 35 Rights, experientially associated with the acquisition of IK. Therefore, true representations of ICBM are Indigenous‐led and deeply rooted in IK and language, cultural values, law, and practices of environmental stewardship associated with the meaningful exercise of Section 35 Rights. In this context, ICBM is not simply monitoring what remains, but also restoring IK through the resurgence of accessible traditional and cultural practices. When assessing impacts on land, there is an equal need for natural scientists to understand community perspectives, methods, indicators, and thresholds and for communities to understand WS methods of data collection and analysis. This report highlights key principles to meaningfully guide collaboration between researchers and community members, including the 10 Calls to Action developed by Wong et al.  and refeIn reviewing best practices and principles of existing ICBM and WS programs involving Indigenous peoples in the OSR, focusing on the period 2009–2020, we discuss the importance of ICBM and WS‐based programs involving Indigenous communities in the OSR. Although the idea of integrating IK and WS to tackle challenging environmental problems is not new, the paper provides a unique way of organizing the integration of IK and WS using a conceptual model and a governance structure to support these collaborative efforts. It is not the intention of the present paper to adapt IK or WS to any model or monitoring program, but to discuss the integrative ability of both to better understand pressure/stressor‐pathways and response relationships of environmental impacts in the OSR. Contextualizing each program into the conceptual model is useful for large monitoring programs, like the OSM program, to integrate large amounts of data assessing different aspects of the environment over time and space. Using a conceptual model can enable regional monitoring programs to constantly evaluate research objectives and identify gaps in a collaborative way. As with the OSM program, the conceptual model is meant to be adaptable and inclusive of all knowledge systems and can be interpreted in many ways. Regardless of interpretation, the conceptual model is not complete without the application of both IK and WS.Direct input from communities would have been valuable for this paper; however, other priorities prevented participation beyond internal governance structure. Communities may have their own ways of presenting relationships in the conceptual model; however, the presentation of relationships in this paper still offers the opportunity to examine the possibilities of data integration (WS and IK) to answer complex environmental questions. It is important to note that communities participating in the OSM program focus on reports generated for community use rather than traditional WS outputs. Although this is important for communities, it has limited the availability of reports and peer‐reviewed publications from ICBM initiatives for this review. As a result, a significant amount of the literature included in the review is gray literature or reports collected from communities, organizations, and government websites. As programs mature, more ICBM reports can be made available to the public and for OSM program use.Currently, many ICBM and WS science programs involving Indigenous peoples in the OSM are still in their infancy. Best practices of ICBM were largely represented in ICBM programs led by MCFN and ACFN, most which were initiated before the formation of the OSM program. As leaders in ICBM, the MCFN and ACFN play a large role in the OSM program. ICBM initiatives by the ACFN and MCFN, although holistic in practice, focus predominantly on the impacts of contaminants on air and water. Currently, there is less documentation of land (including wetlands) and terrestrial wildlife monitoring. However, this does not characterize land and wildlife as less significant, but suggests that ICBM approaches currently support regional monitoring efforts where data are more easily captured in regional‐scale models, such as air and water quality. Data repositories and data mapping tools sensitive to the collection of various IK  are gaps in the OSM program that require attention. Valuable integration opportunities for ICBM exist in the OSM program, specifically with respect to aligning or validating geospatial products with community‐based observations of near‐real‐time environmental change.Overall, from a regional perspective, communities can benefit from working with other Indigenous communities and scientists to share knowledge and data when participating in monitoring programs. Increasing the scope (area) of data collection can help address issues of incorporating local data into regional‐scale models. Using this strategy, communities can discuss what they want to know (research questions), what will be monitored (when and where), and how strategies integrating IK and WS can improve environmental monitoring over a large area. Regional‐scale data can improve the power of ICBM data in regional monitoring programs, advancing the ability to detect the likelihood of environmental impacts over time and space. In all capacities, the involvement of Indigenous peoples in the OSM program has moved the program closer to achieving this goal. Opportunities exist to increase data collection over time and space and, potentially, in areas not easily accessible by non‐Indigenous peoples. This can increase the potential for regional monitoring programs to address community concerns and also fill relational gaps visually guided through the use of conceptual models. When developed appropriately, ICBM programs can enhance the effectiveness and relevancy of environmental monitoring linking physical and chemical stressors (land disturbance and contamination) with culturally significant indicators  and address community concerns.The authors declare no conflicts of interest.This article is written through a Western science lens, and the knowledge from, and perspectives of, Indigenous communities are largely absent. Communities were consulted through the governance of the OSM program; however, the development of ICBM framework through the ICBM advisory committee was a priority during this time. Information from Indigenous participants undertaking, collaborating, and/or participating in Indigenous community‐based monitoring programs are important components of the development of ICBM initiatives. Although the authors of this paper recognize the inextricable link between environmental health and human health, human health was outside the scope of this environmental monitoring review. Through an Indigenous lens, this framework is incomplete without the addition of human‐health considerations.Supplemental Table S1 is associated with a conceptual model in the paper , which organizes literature along a stressor‐pathway‐response model.This article includes online‐only Supporting Information.Supporting information.Click here for additional data file.


The RIKEN Nishina Center (RNC) executed an accelerator upgrade project for the heavy-ion linac . A superconducting RIKEN linear accelerator (SRILAC) and a new superconducting electron-cyclotron-resonance ion source (SC-ECRIS) to boost the final energy and intensity were constructed, aimed at synthesizing a new superheavy element, 119, through a hot fusion reaction. The project included the construction of a gas-filled recoil ion separator (GARIS-III) suitable for detecting the residues of the hot-fusion reaction. To avoid research interruption during the SRILAC construction period (2017–2019) and gain experience in hot-fusion reaction processes, GARIS-II located in the GARIS experimental hall in LINAC building was moved to the E6 experimental hall in Nishina building. Certain exploratory measurements were performed employing the beams accelerated by RILAC2 and the RIKEN ring cyclotron (RRC), which is a part of the existing accelerator complex of the radioactive isotope beam factory (RIBF). Further, commissioning experiments with the upgraded facility (SRILAC and GARIS-III) were performed. The upgrade project and its commissioning results are chronologically described in this article. The RIKEN Nishina center (RNC) was established in 2006 to operate/manage the accelerator complex. The Bird’s eye figure of the present RNC facility where research using various heavy-ion beams is being actively pursued is shown in Fig. RIKEN has a long nuclear-science research tradition pioneered by Yoshio Nishina  who consu was provided by RILAC. The evaporation residue, One of the recent epoch-making achievements of the RNC is the discovery of element nihonium –5. Nihonn and 4n reaction channels can be utilized leading to isotopes, namely, In 2016, the RNC commenced a new comprehensive superheavy element (SHE) research program, abbreviated here as the “SHE project” for convenience. Its main objective was to expand the periodic table of elements by synthesizing new superheavy elements. After the discovery of oganesson Z = 118) [ [8], thementally .Table u  operating at a higher RF frequency to increase the beam current.Thus, the primary focus of the SHE project is the upgrade of the RILAC accelerator by replacing in part a superconducting RIKEN linear accelerator (SRILAC) to increase the final beam energy from 5.5 MeV/GARIS, used for isolating and detecting nihonium events, was designed with a high-transmission efficiency for the recoiling evaporation residue produced in a cold fusion reaction. However, its transmission efficiency is significantly reduced for the evaporation residues produced in hot fusion reactions due to their lower recoil velocity compared to cold fusion reactions. Therefore, a new gas-filled recoil ion separator GARIS-II suitable for hot-fusion reaction products was designed and constructed . It was It was estimated that at least three years would be required for the construction of SRILAC. To avoid research interruption during the SRILAC construction period and gain insights on the hot-fusion reaction, GARIS-II was moved to experimental hall E6 in Nishina building see Fig.  in 2018.The existing accelerators, the injector linac  and the RIKEN Ring Cyclotron (RRC), together with the vital SC-ECRIS rendered certain exploratory measurements possible using an intense The upgrade project concluded with the newly constructed GARIS-III in the experimental hall in LINAC building where GARIS-II was previously located. GARIS-III is essentially a copy of GARIS-II.In 2020, commissioning of the entire system  commenced. SC-ECRIS and SRILAC completely satisfied the planned specifications.The upgraded accelerator system was designed to deliver a mentclass2pt{minimThis contribution describes the SHE project at the RNC, which commenced in 2016, with particular emphasis on the accelerator facility upgrade Sect. . Furthered Sect. , followeu at 37.75 MHz as an injector for the RIBF accelerator complex [The original RILAC  comprise complex .Fig. 2Ovu) [u to 5.5 MeV/u by adding a booster linac called charge-state-multiplier (CSM) system [Element Nh was synthesized by bombarding a entclass1pt{minimant}μA u) . The ene) system  comprisiu is required, necessitating energy upgrade. Therefore, a superconducting-linac (SC-linac) was proposed [u to u. Given the limited space in the existing RILAC building, SRILAC was installed by replacing four CSMs  was also studied for the RIBF to gain more beam power up to 11 MeV/ium beam , 34.The schematic of the SC-quarter-wavelength resonator (SC-QWR), based on this SRILAC design, is shown in Fig. U is the stored energy, SC-QWRs are made from highly purified Nb sheets. The bulk Nb cavity consists of an outer-cylinder, a stem, and top- and bottom- end caps welded through electron-beam-welding. After welding, the inner surfaces are treated by a standard processing method based on buffered chemical polishing and high-pressure rinsing with ultrapure water. Before installing a local magnetic shield  and a medium-energy beam-transport-line (MEBT), which connects the CMs, RILAC DTLs, and the high-energy beam transport line. Both CM1 and CM2 contain four SC-QWRs each. The schematic of CM1 and CM2 is shown in Fig. The design of CM3 is the same as that of CM1 and CM2 but it contains only two SC-QWRs. Instead of the traditional top-loading type, bottom-up-type CMs are used. The 4K cold mass, which comprises a cavity, a Mu-metal local magnetic shield, a helium vessel, FPC, and a dynamic tuner , pressure change of the supplied helium, and Lorentz force detuning.The function of MEBT is to maintain the vacuum, transversely focus the beam, and perform beam diagnostics.For beam transport, room-temperature quadrupole magnets, which include horizontal and vertical steering functions, are used.Instead of traditional beam diagnostic devices, such as wire scanners and Faraday cups, the beam-energy-position monitor, named BEPM, is employed. The BEPM is intended to simultaneously measure the timing, position, energy, longitudinal profile, and amplitude of the beam. Eight BEPMs are installed to measure the beam properties before and after acceleration by each CM. The BEPM for SRILAC was developed along with the Beam Diagnostics Group of J-PARC .As beam measurement is nondestructive, ideally, there is neither outgassing nor spattering to produce dust, which degrade the field emission on the superconducting cavity surface.One of the most critical issues in the design of the beam transport line is the prevention of SC-cavity contamination by the dust transported from the room-temperature section through gas flow due to the vacuum pressure gradient. While the vacuum-pressure level of the SC part drops as low as 1 eveloped . This diThe new SC-ECRIS is structured to generate a powerful magnetic mirror for confining the extremely hot electrons heated by high-power microwaves. As shown in the schematic Fig. , the RIKIn addition, using the six SC solenoidal coils, it is possible to generate various mirror field distributions along the ion-source axis, for example, either “classical nt}Bmin” . This mant}Bmin” .For long-term operation at the RIBF, the ion-source operation is optimized to minimize the X-ray heat load, which destroys the insulation materials in the cryostat chamber.To provide metal ions such as uranium or vanadium for a period of one month, an ion-source structure with a pair of high-temperature ovens was developed and installed . An inteAfter the hardware was commissioned, in January 2020, the beam acceleration test was conducted for the first time. The specifications of SRILAC are listed in Table u with a gap voltage of 1.13 MV/cavity. For the acceleration test, only nine SC-cavities were used because there was a vacuum leak with the ceramic window of the FPC of cavity No. 5 (SC05) then. For SC-linac tuning, the SC-QWRs were energized one-by-one, and the beam energy was measured, with the systematic variation of the RF-field phase  method using a pair of BEPMs with low-beam current. The RF phase of each cavity must be synchronized with the timing of the beam bunches. Then, the energy gain of the beams passing through the cavity can be denoted as The vertical and horizontal beam positions monitored by the BEPMs were finely centered; the beam loss, which mainly occurred in MEBT, was minimized, keeping the deterioration of the vacuum pressure below 1u was accelerated and delivered. Stable beams with the required energy and intensity were successfully provided after careful tuning, and a beam power of more than 1 kW was achieved.After beam commissioning and the mitigation of various problems, user beam service commenced. A u, which is the optimal energy for producing From the accelerating structure point-of-view, one of the advantages of the double-gap cavity, adopted in the SC-QWR for SRILAC, is the high-flexibility in accelerating the beam energy compared to multicell drift tube linacs. Utilizing the independently phased array of the QWR, the accelerating beam energy can be set within a wide range and finely tuned by selecting the number of cavities and gap voltages. For example, the acceleration of light-ions, such as alpha particles, is possible at 7.25 MeV/Thus far, a GARIS-II in the RILAC experimental hall in the LINAC building, which had been operational since 2013, was transferred in 2018 to the E6 experimental hall in Nishina building together with the gas-cooled rotating target system, differential pumping system, FPD system, and analog electronics.At the new location in E6, the available area for GARIS-II was limited by the floor and wall structures. Therefore, the GARIS-II magnets were reversed to change the ion deflection angle from left to right for fitting into the available floor space. Dedicated beam diagnostics for GARIS-II were newly installed.One of the booster linac CSMs used in RILAC, which was decommissioned for adopting to SRILAC, was transported in the beam transport line of GARIS-II. It was used for modifying a small quantity of beam energy, such as ± 3%. In addition, a ToF system was installed to measure the beam velocity. The entire relocation process required half a year for completion.u for the SHE experiments. However, in the standard RRC operation for RIBF experiments, ions are accelerated to an energy of 10.7 MeV/u by the combined operation of RILAC2 and RRC at an RF frequency of 18.25 MHz and harmonic number (H) nine. To match a specific energy of u for the SHE experiments, H = 12 was selected. One of the operational issues for H = 12 is that the acceleration voltage of the RF cavity (frequency-tunable double gap type) is as low as 60 kV/gap at an RF frequency of 18.25 MHz. This acceleration voltage is insufficient for handling high-power beams with the space charge effect. To solve this problem, the two double-gap resonators [In parallel with the relocation, the RRC was modified to accelerate sonators  were remBy 2018, SHE experiments could commence using the accelerator complex, 28-GHz ECRIS, RILAC2 and RRC, and the transferred GARIS-II system.By the time SRILAC was completed, GARIS-III was constructed and installed in the RILAC experimental hall to conclude the main part of the SHE project. Note that GARIS-III is a duplication of GARIS-II and is placed in the previous location of GARIS-II next to GARIS. The magnet configuration for both GARIS-II and GARIS-III includes two dipole magnets (D1 and D2) and three quadrupole magnets  as shown in Fig. However, a minor modification was made based on the operational experience of GARIS-II for reducing the background events at the focal plane. These background events are partially caused by rescattering from the inside surface of the D1 vacuum chamber. A rectangular  at each focal plane. It was found that the background count rate of GARIS-III was reduced by The design, construction, and performance of GARIS-II are reported in Ref. . The ionnt}283Cn . AccordiOnly the new aspects associated with the installation of GARIS-III are described below.Figure The inset photograph in Fig.  curtain , target  curtain , 45. TheBetween the exit of SRILAC (CM3) and the 90ectively . This enIn the DPS, seven turbomolecular pumps (TMP) with an evacuation speed of 350 l/s and a mechanical booster pump (MBP) with an exhaust rate of 280 l/s are employed. In addition, an MBP with an exhaust rate of 280 l/s is attached to the Because the differential pumping system has a poor pumping speed for He gas, a device called ‘The Charge-state variation of the evaporation residues was not observed in GARIS-III after the installation of the For the transfer of GARIS-II mentioned in Sec. A Faraday cup can be inserted at the target position to measure the beam intensity when necessary. The beam spot size is confirmed by observing the luminescence of the alumina fluorescent plate  photodiode (Hamamatsu S1223) mounted at 45At the entrance of the target chamber, there is a Ta slit with an aperture sized 11 mmThe FPD system depicted in Fig. The three blue circles shown in the figure are 0.5 A newly developed ToF detector was installed and used in the FPD of GARIS-II located in the RILAC experimental hall, as shown in Fig. 1 of Ref.  before rThe aperture of the ToF detector used in GARIS-II was increased compared to that used in GARIS, from 57 cmreaction . The perThe Si detector box contains two double-sided silicon strip detectors (DSSDs) for implantation surrounded by a tunnel detector, and a Veto detector located behind the DSSD. Each DSSD with a thickness of 500 AT Inc.) . The tunTwo DSSDs arranged side-by-side cover the necessary sizes of the focal plane image  was measured using the ly known . The reaGARIS-II .xn) Further, the excitation functions of the ARIS-III . The rean to 4n evaporation channels are plotted in Fig. The results of the excitation functions for the 2The obtained results for the RV) code  and the RV) code . The staAfter the completion of GARIS-II, a commissioning experiment to synthesize element 118, oganesson, through hot fusion reaction building . A \docuHz ECRIS  through Hz ECRIS . The resHz ECRIS .Curium in an oxide form , and was subsequently continued with the newly built SRILAC and GARIS-III facility.Unfortunately, as the commissioning and search for a new element were frequently interrupted during 2020 and 2021, the efficiency was below expectation. The various delays caused by the COVID-19 pandemic is an apparent reason. Moreover, a severe breakdown of the 40-year-old linac, RILAC, occurred. It should also be mentioned that the highly efficient Efforts are being made to accumulate a dose equivalent to t}∼10 fb , 15–17, In principle, the RNC can currently execute two new SHE search experiments simultaneously with the SRILAC +GARIS-III facility and the RILAC2+RRC+GARIS-II facility if sufficient manpower, target material, and beam time are available.However, the RRC accelerator is extensively used for RIBF experiments. Thus, obtaining beam time for the SHE search experiment with GARIS-II, which requires considerable of beam time, is challenging. Nonetheless, GARIS-II is highly attractive for research that requires relatively short beam time. For example, the first direct measurement of the atomic mass of superheavy nucleus al plane .In the SRILAC+GARIS-III facility, the available beam time can be utilized by the new element research group for measurement with an intense


No characteristic dominant genus was observed across the IGM samples. The IGM pus samples were more diverse than corresponding IGM skin samples . Corynebacterium kroppenstedtii, reportedly associated with IGM in the literature, was higher in IGM pus samples than paired skin samples . Three other species and nineteen genera were statistically significant in paired IGM pus–skin comparison after antibiotic treatment adjustment and multiple comparisons correction. Microbial profiles are unique between patients with IGM and LM. Inter-patient variability and polymicrobial IGM pus samples cannot implicate specific genus or species as an infectious cause for IGM.Idiopathic granulomatous mastitis (IGM) is a rare and benign inflammatory breast disease with ambiguous aetiology. Contrastingly, lactational mastitis (LM) is commonly diagnosed in breastfeeding women. To investigate IGM aetiology, we profiled the microbial flora of pus and skin in patients with IGM and LM. A total of 26 patients with IGM and 6 patients with LM were included in the study. The 16S rRNA sequencing libraries were constructed from 16S rRNA gene amplified from total DNA extracted from pus and skin swabs in patients with IGM and LM controls. Constructed libraries were multiplexed and paired-end sequenced on HiSeq4000. Metagenomic analysis was conducted using modified microbiome abundance analysis suite customised R-resource for paired pus and skin samples. Microbiome multivariable association analyses were performed using linear models. A total of 21 IGM and 3 LM paired pus and skin samples underwent metagenomic analysis. Bray−Curtis ecological dissimilarity distance showed dissimilarity across four sample types (IGM pus, IGM skin, LM pus, and LM skin; PERMANOVA,  Idiopathic granulomatous mastitis (IGM) has been consistently described as a rare and chronic inflammatory infliction of the breast ,2. The cCorynebacterium was first associated with IGM when Taylor et al. (2003) reported coryneform bacteria observed in histological examination of breast tissue and Corynebacterium species isolated in cultures from patient specimens .,51.Ochroue cases . Furtherue cases . Ochrobaas well) . Howeverettings) , whereasStreptococcus and Staphylococcus infections, with Erythromycin targeting Gram-positive bacteria that are often implicated in granulomatous inflammations [Patients with IGM self-reported treatment with Cephalexin, Amoxicillin-clavulanate, Clindamycin, Trimethoprim-sulfamethoxazole, and/or Erythromycin largely targets mmations ,55,56,57To the best of our knowledge, this is the first study for patients with IGM that provides the duality of intra-patient controls with paired skin and pus samples, on top of the comparison with patients with LM. Furthermore, the use of 16S sequencing is less common for IGM studies, beyond bacteria culture methods, so as to profile both living and dead microbial populations. Metagenomic analysis matching patient samples to previous antibiotic treatment would also identify antibiotic-resistant strains. However, as a rare condition, the number of patients with IGM recruited is limited. The limited LM control patients, as well, means comparisons and conclusions made on the basis of imbalances between IGM and LM patient numbers in our study must be validated in a larger cohort. Future skin sample collections must control for washing and exposure (cloth covering) before sample collection. Subsequent studies should also use swabs moistened with preservation medium or enzymatic lysis buffer, as opposed to dry swabs, for skin sample collection, for improved biomass collection . It mustthird generation sequencing platforms to sequence the entire 16S rRNA gene ; orshot-gun metagenomics for sequencing the entire microbial genome .The study population was adult patients (aged 21 years and above) suspected, diagnosed with or treated for mastitis (IGM or LM) at five participating hospitals in Singapore: National University Hospital (NUH), Tan Tock Seng Hospital (TTSH), KK Women’s and Children’s Hospital (KKH), Singapore General Hospital (SGH), and Changi General Hospital (CGH). Pregnant and breastfeeding women, and lactating and non-lactating women were allowed to participate in this study. Patients who had received or were receiving treatment at the point of recruitment were allowed to participate in the study. Patient information on demographic, lifestyle, reproductive, past and current treatment or environmental exposure variables were collected from questionnaires. All studies were performed in accordance with the Declaration of Helsinki. All participants provided written informed consent. This study was approved by the National Healthcare Group Domain Specific Review Board (reference number: 2017/01057) and the Agency for Science, Technology and Research Institutional Review Board (reference number: 2020-152).Adult female patients with IGM or LM were recruited from the five participating hospitals in Singapore between 2018 and 2020. Target recruitment was originally 60 participants per site, totalling 300 participants in the study. During the recruitment time period, 148 patients were screened, 91 patients enrolled in the study, and 89 patients completed the study . The main study recruited patients who donated any of the following: blood, saliva, tissue, pus swab, and / or skin swab. In this metagenomic study, a subset of 32 patients provided pus swabs, and / or skin swabs, of which 24 were included for metagenomic analysis .Mycobacterium tuberculosis infection, and negative stains for Grocott (methenamine) silver (GMS) stain or Periodic acid–Schiff (PAS) to rule out fungal infections. Histopathology also confirmed non-malignancy. Patients presenting with mastitis and/or breast abscess confirmed to be lactating at the point of recruitment were diagnosed as LM.Study coordinators were informed by clinicians and physicians of potential mastitis cases, after which, study coordinators would seek consent, collect patient samples and conduct the questionnaire. Patients were subsequently followed up for clinical diagnosis confirmation. Patients who were not subsequently diagnosed as IGM or LM were excluded from the study. Non-lactating patients presenting with mastitis and/or breast abscess were diagnosed as IGM by histological examination of breast core biopsy for non-caseating granulomatous formation. Other conditions were excluded via negative stains for acid-fast bacillus (AFB) to rule out Patient samples were collected using DNA/RNA Shield Lysis Tubes w/Swab (Microbe) . Sterile methods were used to collect pus samples from the infected breast, while skin samples were collected from the contralateral (non-infected breast) without site cleansing prior to sample collection. The kit sample collection tubes, that were pre-filled with DNA/RNA Shield and a lysis reagent for microbial samples, were labelled before sample collection. Swab was removed from packaging without touching the applicator tip.Pus sample collection: Swab was allowed to absorb infected fluid for 30 s while rotating the swab;Skin sample collection: Swab was used to rub skin area for 10 to 15 strokes with moderate pressure. Swab was rotated and sampling was repeated.Swab was broken along the breakpoint and the applicator head was left in the collection tube. Each collection tube was capped and shaken 10 times to mix the sample collected with pre-filled reagents. Total DNA was extracted with ZymoBIOMICS™ 96 MagBead DNA Kit . Samples were mechanically lysed with beads beating at maximum speed for 20 minutes. DNA was purified from lysed samples with magnetic beads (ZymoBIOMICS™ MagBinding Beads) and eluted with ZymoBIOMICS™ DNase/RNase Free Water. The 16S ribosomal RNA (rRNA) gene amplification and sequencing has been previously described . BrieflyDenaturation at 95 °C for 30 seconds;Annealing at 59 °C for 30 seconds; andExtension at 72 °C for 1 minute.® Ultra™ II DNA Library Prep Modules for Illumina®  according to the manufacturer’s protocol. Multiplexing Sample Preparation Oligonucleotide Kit from Illumina, Inc.  was used to label the DNA sequencing libraries with different multiplex indexing barcodes [The final elongation was at 72 °C for 6 minutes. QIAquick PCR Purification Kit  was used to purify 700–1000 bases PCR products. Amplicons were sheared with Covaris Adaptive Focused Acoustics™ to approximately 180 base fragments. The 16S libraries were prepared using NEBNextbarcodes . The 200https://greengenes.secondgenome.com/?prefix=downloads/greengenes_database/gg_13_5/, accessed on 19 November 2020) using Basic Local Alignment Search Tool (BLAST) [Processing of sequencing data has been previously described . Briefly (BLAST) . The top (BLAST) . Percenthttps://github.com/BetaCollins/MBAA-Suite, accessed on 16 December 2020) customized R resource that contains the analysis functions of 16S pipeline  developed at the Genome Institute of Singapore, designed for Illumina shotgun sequencing of 16S rRNA amplicon sequences [Statistical analysis was performed using a modified microbiome abundance analysis-suite ) and Simpson (Equation (2)) indices were used to measure alpha-diversity.Corynebacterium genera was evaluated in pus and skin samples from patients with IGM and LM. Paired sample analysis with Wilcoxon signed-rank test, compared Corynebacterium genera relative abundance in pus and skin samples from the same patient in patients with IGM. Microbiome multivariable association was performed with linear models using MaAsLin 2  [Multidimensional scaling was used for visualisation of Bray−Curtis distance between the microbial profiles of the IGM and LM sample types. Permutational multivariate analysis of variance (PERMANOVA) was used to make comparisons of mastitis and sample types centroids and dispersion within groups; statistical significance was defined by measure of space not equivalent for all groups. Relative abundance of ch 2022)  to compaAll analyses were performed with R (v4.0.2) unless otherwise stated. Datasets utilised in analysis can be made available from the corresponding author upon reasonable request, within limitations of the study Institutional Review Board (IRB).Corynebacterium genus was not significantly different between paired pus and skin samples, the Corynebacterium kroppenstedtii species was, and so were 19 genera and 3 other species. However, the inter-patient variability and polymicrobial microbiome of IGM pus samples cannot implicate specific genus or species of bacteria in an infectious cause for IGM. Subsequent investigations with improved sampling methods, and employment of advanced metagenomics methods can achieve higher resolution for categorical definition of microbial populations in IGM pus and skin samples. Studies with larger patient numbers, backed by molecular experiments, are needed to confirm or discard an infectious aetiology for IGM.Microbial profiles are unique between patients with IGM and LM. Our study has demonstrated statistically significant difference in the microbial communities of IGM pus, IGM skin, LM pus, and LM skin samples. While