BackgroundAlthough considerable research on physician fellowship training program websites has demonstrated critical deficiencies in accessibility and content, there is no published study regarding the website content of physician assistant/associate (PA) and nurse practitioner (NP) orthopedic surgery postgraduate residency/fellowship programs in the United States. Therefore, this study aims to conduct a web-based content analysis to assess the available information on program websites and identify the potential areas for improvement.MethodsThe methodology for this study was replicated from prior research on graduate medical education (GME) websites. Twenty-two PA and joint PA/NP orthopedic surgery postgraduate residency/fellowship training program websites were assessed between August and September 2022. The criteria comprised 17 items related to postgraduate education and recruitment content. All program websites were evaluated for pertinent content relevant to prospective applicants applying to orthopedic surgery residency/fellowship programs. Descriptive statistics were performed.ResultsOut of the 22 PA and joint PA/NP orthopedic postgraduate websites evaluated, all had a functional website link. Only one orthopedic surgery postgraduate website met all 17 criteria. All program websites assessed contained program descriptions and contact information. Other information present on websites included program admission requirements 95% (21/22), salary/benefits 77% (17/22), interview requirement 73% (16/22), faculty listing 68% (15/22), journal club 59% (13/22), program objectives/goals 59% (13/22), and acceptance process 54% (12/22). Less than half of the program websites contained the following information: listing of current PA/NP postgraduate trainees, sample rotation schedule, orientation activities, curriculum, wellness/well-being strategy, graduation and research requirements, and link to program handbook. However, accredited PA orthopedic surgery program websites contained more content relevant to prospective applicants than non-accredited programs.ConclusionThis is the first study to provide a detailed analysis of PA and joint PA/NP postgraduate orthopedic surgery residency/fellowship program websites. Not surprisingly, there is a wide variation in the available content on these websites. Study results provide further evidence to support the urgent need to improve program website content for prospective applicants. Physician assistants/associates (PAs) and nurse practitioners (NPs) often referred to as advanced practice providers (APPs) have been a linchpin of the American Health Care system since the 1960s. PAs are educated as generalists in the medical model, whereas NPs are educated in at least one of six population foci . The groPostgraduate specialty training opportunities are designed to provide greater depth in a specialty or subspecialty area not covered in an entry-level PA and NP education program. Additionally, newly minted PAs and NPs may find these programs attractive because of the opportunity to hone their clinical skills or acquire new ones. For example, Hart and Bowen surveyed several hundred licensed NPs and reported that 90% of them felt either “well prepared” or “somewhat prepared” for practice upon completion of their NP program and expressed interest in transition-to-practice opportunities through a formalized training program such as a residency ,10-13, tCurrently, there is no national resident matching program or centralized application-processing service available to licensed PAs and NPs interested in pursuing postgraduate residency/fellowship training . TherefoA comprehensive list of PA and joint PA/NP postgraduate orthopedic surgery residency/fellowship programs was compiled from the following sources: the Association of Post Graduate Advanced Practice Registered Nurse (APRN) Programs (APGAP), the Association of Postgraduate PA Programs (APPAP) websites, a Google search strategy, and from prior research on PA and NP postgraduate orthopedic surgery programs . The GooOther external sources of information outside of the program’s dedicated website were not examined due to the potential for outdated and/or incomplete information. Each program's website home page was scanned along with content searches within each website to identify the presence of 17 criteria based on two categories. The postgraduate recruitment category included a listing of faculty and current fellows/residents, admission requirements, program description, acceptance process, salary/benefits, contact information, interview requirement, and program objectives/goals. The postgraduate education category included journal club, sample rotation schedule, orthopedic curriculum, orientation activities, graduation and research requirements, resident wellness/well-being, and hyperlink to program handbook. All 17 criteria were weighted appropriately. For example, when a content item was found on the program’s website homepage and/or subpages, it was awarded 1 point, but if absent, it was awarded 0 points. The rationale for including certain criteria elements in this study is based on the findings from previous research. For instance, in one study, medical students applying to surgical specialties indicated that fellowship acquisition, faculty information, application contact information, and resident wellness are the most important website content . AnotherThis study does not require institutional review board approval since the information assessed is from publicly available websites and there was no contact with residency/fellowship programs. The 22 PA and joint PA/NP postgraduate orthopedic surgery residency/fellowship programs are located in seven specific US census regions/divisions , salary/benefits 77% (17/22), interview requirement 73% (16/22), faculty listing 68% (15/22), journal club 59% (13/22), program objectives/goals 59% (13/22), and acceptance process 54% (12/22). Less than half of the program websites contained the following information: listing of current PA/NP postgraduate trainees, sample rotation schedule, orientation activities, curriculum, wellness/well-being strategy, graduation and research requirements, and link to program handbook  accredited PA orthopedic surgery program websites contained more content criteria relevant to prospective applicants than non-accredited programs. Not surprising, given that external accreditation requires strict compliance with administrative and educational standards. Nevertheless, PA and NP postgraduate residency/fellowship specialty programs are not required to seek accreditation. Although not part of the original criteria, four websites (18%) included online video content, which offered a compelling view of postgraduate specialty training.This is a rare study to examine the accessibility and presence of website content pertinent to PA and NP orthopedic surgery residency/fellowship applicants. Usability testing and quality assessment of the information provided on program websites were not conducted. While the results of this study indicate that the most prevalent information on program websites were program descriptions, contact information, salary/benefits, and interview requirement, there were substantial gaps in education and recruitment content. Information details about wellness, graduation requirements, sample rotation schedule, curriculum, link to program handbook, and listing of current residents/fellows were least readily available. These important findings coupled with existing research on GME programs highlight the value of enhancing program website content, which may improve orthopedic surgery recruitment efforts ,18. MoreA call to actionOur world today is undeniably digital, where data, people, and devices converge. Information is readily available and highly sought after by prospective applicants. Therefore, postgraduate programs should regularly audit their website content to ensure their online information is easily accessible, loads quickly, and is up to date. As an immediate step, orthopedic surgery postgraduate programs should clearly articulate on their website for prospective applicants the minimum graduation requirements and essential knowledge, skills, and competencies that must be satisfied to earn the certificate of completion from the program. Also, program websites should include a listing of current fellows/residents along with faculty photos and biography that present faculty members' education and research interests to prospective applicants. This is underscored by a retrospective analysis using website analytics of an accredited emergency medicine residency training program, which found that the most popular content accessed was the “people” section of the website, which includes photos of current residents and faculty .Additionally, postgraduate programs should display an example of a weekly rotation and call schedule on their websites for prospective applicants to help codify training expectations. This is extremely important as some applicants prefer lifestyle controllability over traditional extended shifts as burnout rates are reportedly higher among those undergoing overnight call rotations . FurtherLimitationsThis study is not without limitations. First, data elements were initially completed by a single researcher on two separate occasions, which may have led to some degree of bias. To reduce bias, an independent reviewer blinded to the results of the first review extracted data on 12 websites (54.5%) to analyze the online search results. Any data collection discrepancies led to the author and independent reviewer reexamining the specific website to resolve any errors. Additionally, this study was conducted on websites from a single specialty, thus limiting generalizability to other PA and NP postgraduate specialties. Moreover, the content analysis did not account for any program website updates or development of any new orthopedic surgery residency/fellowship programs that occurred after the review period. Finally, 17 content items were chosen for this study, but there may have been other content items that PA and NP applicants also find important when considering whether to pursue postgraduate training such as facilities information, housing/neighborhood information, postgraduate job prospects, diversity statement, moonlighting opportunities outside of the program, and loan deferment options. Website recruitment and education content appear uneven across orthopedic surgery postgraduate programs. Given the variation in the online program information, there is an urgent need to optimize the program website content for prospective PA and NP orthopedic surgery applicants. Future research is needed to identify whether specific website content influences the decision of PA and NP applicants to apply to a postgraduate program. Moreover, whether prospective applicants search for residency/fellowship program information from online resources offered by APPAP or APGAP is an area of emerging interest. Also, an investigation into the quality, usability, and accessibility of PA and NP postgraduate residency/fellowship website content across all specialties is warranted.


This study was conducted to develop a couple-centered antenatal education program and to test the program’s feasibility. With a preliminary-experimental study design, 33 pregnant couples who were expecting their first child participated in this study. The program consisted of four sessions (1 hour/session/week) of education and counseling. Data were collected before and after the intervention from September 2018 to April 2019 at a women’s hospital in Daejeon, Korea, with demographic data forms, the Edinburgh Postnatal Depression Scale, Perceived Stress Scale, Maternal–Fetal Attachment Scale, Korean Newborn Care Confidence Scale, Wijma Delivery Expectancy/Experience Questionnaire, and Dyadic Adjustment Scale-10. The pregnant women and their husbands were on average 32.30±3.10 and 33.21±6.25 years old, respectively. The mean marriage duration was 2.34±1.63 years, the gestational age was 31.30±2.66 weeks, and 78.8% of the couples had a planned pregnancy. After the program, both the pregnant women and their husbands showed significant improvements in attachment to the fetus and confidence in providing infant care. Prenatal depression, prenatal stress, and fear of childbirth in pregnant women significantly decreased after completing the program. However, the dyadic adjustment score did not change significantly either in the pregnant women or their husbands.A couple-centered antenatal education program seems to be effective for couples adjusting to parenthood, but further studies should explore ways to have a positive impact on couples’ relationships. Parenthood begins with having the first child; pregnancy is a time of preparation, and the postpartum period is a time of change in the marital relationship due to the new parental role . AnxietyWorsening of the marital relationship has a negative effect on attachment to the fetus, as well as the process of adapting to changes in the marital relationship during the transition period . ConversMercer  theorizeMost prenatal education in South Korea  is provided for only pregnant women (64.4%), with a focus on delivery, breastfeeding, and infant care; meanwhile, very few studies have explored interventions dealing with mother–father interactions or couple relationships , leavingThe hypotheses of this study were as follows.1) Pregnant couples who participate in the couple-centered antenatal education program will have a better adaptation to becoming parents, as shown by measures including antenatal depression, antenatal stress, fetal attachment, newborn-care confidence, and mother’s fear of childbirth.2) Pregnant couples who participate in the couple-centered antenatal education program will improve their adaptation to the couple relationship, as measured by the couple’s dyadic adjustment.Ethics statement: This study approved was by the Institutional Review Board of Chungnam National University (201806-SB-063-01). Informed consent was obtained from the participants.The couple-centered antenatal program was developed by the first author through multiple stages of program development, as shown in The study participants were married couples, including women who were pregnant with their first child at 28-34 weeks of gestation, because couples who answered that they were well prepared in the third trimester were found to adapt well in the postpartum period . Since tThe required sample size was calculated based on effect sizes (d) of 1.71 and 0.47 for a childbirth education pilot study aimed at preventing fear of childbirth and depression, respectively, among pregnant women . For theA couple-centered antenatal education program was developed to help couples’ adaptation to become parents and their relationship. The environmental interaction model  of the pThe couple-centered antenatal education program applied in this study  had two Two nurses  provided the educational program in four sessions, each of which lasted for 1 hour in the childbirth education room of the hospital. The size of each intervention group was decided based on couples’ needs, with the education provided to one to three couples at a time. The teaching method consisted of face-to-face education and counseling. Throughout the session, we provided education and information to help the couples discuss these topics, allocated time to practice skills such as expressing one’s own emotions and effective conversation techniques, and provided time for questions and answers. In addition, a small gift was presented to encourage participants to continue participating in the program, and periodic text messages were sent as reminders before scheduled appointments and to express appreciation for the participants’ participation.The study variables were measured before and after the educational program to test the program’s effectiveness. The researcher explained to the couple how to answer each questionnaire and asked them to respond separately to prevent couple-related dynamics from affecting the responses. The measures for adaptation to becoming parents were antenatal depression and stress for the parents themselves, parental-fetal attachment for the parental-fetal relationship, newborn-care confidence for becoming parents, and fear of childbirth for mother’s childbirth preparation, while dyadic adjustment was used to measure adaptation of the couple relationship.Depression in pregnant couples was measured using the Korean version of the Edinburgh Postnatal Depression Scale , which wStress in pregnant couples was measured using the Korean version of the Perceived Stress Scale , which wParental-fetal attachment was measured using the Korean version of the Maternal–Fetal Attachment Scale , which wConfidence in caring for newborns was measured using a self-report questionnaire . The 22-Childbirth fear in pregnant women was measured using the Korean version of the Wijma Delivery Expectancy Questionnaire (version A)  before dMarital adaptability was measured using the Dyadic Adjustment Scale (DAS)-10 , which iInformation was gathered on participants’ age, educational background, occupation, marriage duration, family income, whether the pregnancy was planned, gestational age, and lactation plan.Participants were asked about their satisfaction with the overall program content, the topic that they most enjoyed among the four sessions of the program, any material that they thought was missing or could have been supplemented, perceptions regarding any other educational content that would be needed in pregnancy and childbirth education, and the educational content that they desired in postnatal education programs.The effect of the intervention in a single group was analyzed using the one-tailed paired t-test using IBM SPSS ver. 25.0 .The pregnant women were 32.30±3.10 years of age (mean±standard deviation), and their husbands were 33.21±6.25 years of age. Almost all of the pregnant women (97.0%) and all of their husbands had at least a college education, 72.7% of the pregnant women and all of their husbands were employed, and 42.4% of the families had a monthly family income of 4–5 million Korean won . The average marriage duration was 2.34±1.63 years, the average gestational age at the time of recruitment was 31.30±2.66 weeks, and 78.8% of the participants had a planned pregnancy .p=.034). The score of the husbands decreased from 3.73±2.52 to 3.67±2.88, but this change was not significant . The stress score of the pregnant women decreased significantly from 15.30±5.48 to 13.33±5.75 , while their husbands showed a non-significant decrease from 12.70±4.66 to 12.27±5.05 .The score for antenatal depression of the pregnant women decreased significantly from 6.12±3.67 before the intervention to 5.18±3.62 after the intervention , as did that for paternal-fetal attachment, from 79.85±12.18 to 84.30±10.02 .The score for maternal-fetal attachment increased significantly from 73.30±13.37 to 77.67±12.86  and from 79.88±17.83 to 83.39±12.78 in their husbands .The score for confidence in providing newborn care increased significantly from 73.27±12.42 to 78.58±12.51 in the pregnant women .Pregnant women’s score for fear of childbirth decreased significantly from 68.97±19.76 before the intervention to 63.45±20.01 after the intervention . Their husbands’ scores decreased by 0.67±3.81, from 43.88±3.28 before the intervention to 43.21±4.64 after the intervention, but this change was likewise not statistically significant   .Over the four sessions, two of the 33 couples were absent once, due to the difficulty of moving to another city and the poor condition of the pregnant woman on the day of the session. Educational videos were provided to the couples who were absent, and after watching they gave their impressions as well as their answers to questions. The program completion rate was 100% when evaluated based on the pregnant couples participating in three of the four sessions, corresponding to a 78% attendance criterion.After completing the antenatal education program, 50.0% of the participants were very satisfied, and 41.4% were satisfied. Satisfaction with the educational topics was highest for newborn care (54.8%), followed by preparation for childbirth (25.0%), communication within couples (12.4%), and adaptation to the pregnancy and postpartum periods (7.8%).This study is the first in Korea to apply couple-centered education in mainstream antenatal education to help adaptation to parenthood in terms of both the marital relationship and the parental role. A unique aspect of the program is that the educational content was well-balanced in terms of adaptation to the parental role and interactions between spouses. In addition, the present study contributes to the expansion of nursing theory and the formation of a body of knowledge in nursing science by constructing a theoretical model of becoming parents for pregnant couples based on the theory of becoming a mother and the intervention effects .The content related to parental adaptation dealt with physical, emotional, and relational changes during pregnancy and postpartum, childbirth preparation, and newborn care. Content related to maintaining the couple relationship was also included, since the couple relationship serves as an important psychological basis for the couple to perform the parental role together . This enThe education on couple-relationship adaptation included emotional intimacy, support, respect, empathy, and emotional exchange between partners after pregnancy and childbirth. This program was offered at an appropriate time for couples to participate together. On-site participation was chosen based on evidence that this setting is effective and that participants prefer to interact with educators, have vivid experiences, and get answers to questions immediately ,43. The The effects of this couple-centered antenatal education program were analyzed in terms of variables for confirming parental adaptation: antenatal depression, antenatal transition stress, parental-fetal attachment, newborn-care confidence, and fear of childbirth. Pregnant women who participated in the program showed reductions in antenatal depression, stress, and fear of childbirth, as well as improvements in maternal-fetal attachment and confidence in providing newborn care. Their husbands showed improvements in attachment with the fetus and confidence in providing newborn care. This is consistent with previous reports that pregnant couples who participated in antenatal education showed increased confidence in parental adaptation , and increased communication and cooperation with their husbands to overcome anxiety and stress ,46. In tHowever, there was no effect on marital adaptation. The score of marital adaptation was 43 and 41 for the pregnant women and their husbands, respectively, in this study, compared with a threshold score of 32 reported by Cho et al. , but theThis study only confirmed the antenatal effects of couple-centered education aiming to help participants become parents in a single group, but a recent meta-analysis showed that couple psychoeducation reduces maternal postpartum depression and ameliorates paternal negative affect, as well as leading to overall improvements in couples’ relationship satisfaction . TherefoFuture studies should consider combining online and offline education in order to reduce the discontinuation of participation related to temporal and spatial restrictions. In the present antenatal education program, scores of higher than 90% were found for both parental adaptation in caring for newborns and preparation for childbirth, and also the satisfaction of the couples with their adaptation during This couple-centered antenatal education program based on the model of becoming parents seems to be effective and feasible as a way to promote the adjustment of pregnant couples to parenthood. Antenatal education programs should be implemented with both couple-relationship and parental adaptation elements to help pregnant couples adjust to parenthood. Future studies should address the need for programs to reflect educational demands related to fathers’ preparations for becoming parents and caring for newborns.


This paper describes one healthcare system’s approach to strategically deploying genetic specialists and pharmacists to support the implementation of a precision medicine program. In 2013, Sanford Health initiated the development of a healthcare system-wide precision medicine program. Here, we report the necessary staffing including the genetic counselors, genetic counseling assistants, pharmacists, and geneticists. We examined the administrative and electronic medical records data to summarize genetic referrals over time as well as the uptake and results of an enterprise-wide genetic screening test. Between 2013 and 2020, the number of genetic specialists employed at Sanford Health increased by 190%, from 10.1 full-time equivalents (FTEs) to 29.3 FTEs. Over the same period, referrals from multiple provider types to genetic services increased by 423%, from 1438 referrals to 7517 referrals. Between 2018 and 2020, 11,771 patients received a genetic screening, with 4% identified with potential monogenic medically actionable predisposition (MAP) findings and 95% identified with at least one informative pharmacogenetic result. Of the MAP-positive patients, 85% had completed a session with a genetics provider. A strategic workforce staffing and deployment allowed Sanford Health to manage a new genetic screening program, which prompted a large increase in genetic referrals. This approach can be used as a template for other healthcare systems interested in the development of a precision medicine program. The use of genetic testing in all areas of medicine is growing, including primary care. In general, primary care providers (PCPs) view the integration of genetics into health care as inevitable, and believe genetic testing provides a clinical utility ,2,3. HowEvidence suggests that PCPs can provide appropriate care and patient management  when adeSanford Health is one of the first healthcare systems to implement a system-wide clinical precision medicine program, including in primary care. Here, we describe how Sanford Health strategically utilized staff and other resources to support its precision medicine program and the growing demand for genetic services. This case study provides a model for healthcare systems to consider when establishing precision medicine programs.CYP2C19 genotyping to inform clopidogrel orders. Efforts culminated in 2018 with the launch of the “Sanford Chip” genetic screen  for 51 medications based on the CPIC guideline updates. This genetic screen also identifies potential MAPs, variants in genes associated with conditions such as hereditary cancers and heart conditions referenced by the American College of Medical Genetics and Genomics (ACMG)s V2.0 list [Sanford Health is the nation’s largest non-profit rural healthcare system with four hubs in Bemidji, MN; Bismarck, ND; Fargo ND; and Sioux Falls, SD. In 2013, Sanford Health provided 2.1 million clinic visits with 35 hospitals and 175 clinic locations. By 2020, Sanford Health provided 5.5 million clinic visits with 46 hospitals and 210 clinic locations. Given the global focus towards precision medicine ,25, Sanfc screen , a genet2.0 list  for the 2.0 list .The precision medicine program achieved its vision by coupling health care professionals with genetic expertise in both clinical and laboratory settings with PCPs and specialty providers. Each member of the clinical team plays an important part in the process of offering the genetic screen and return of results . SpecifiIn 2018, Sanford Health created a new position, “Imagenetics specialists”, with responsibilities similar to those of the genetic counseling assistants . These aPharmacists support the precision medicine program and GCs by answering questions, serving as a reference for PGx results, and communicating with patients about results when requested to do so. Pharmacists review all PGx results and place chart notes in the EMR to alert providers when an actionable finding is identified . PharmacCYP2C19 genotyping and the genetic screen to their patients during the pilot phases of those programs. This group of physicians also served as a resource to colleagues to answer questions. Similar to the physician champions for providers, the PGx pharmacy team identified pharmacist champions who served as resources for a PGx implementation at their individual sites.When the initiative first launched, the four hubs selected “physician champions” who were practicing internal medicine physicians with an enthusiasm about the precision of medicine program. They provided clinical advice about the implementation as members of the precision medicine program steering committee. They were also among the first providers to introduce preemptive The molecular laboratory, which employed a part-time GC before the precision medicine program launched, hired a second laboratory-based GC to support the precision medicine program. The laboratory GCs created standard operating procedures for processing the genetic screening results and contributing to the development of educational materials for providers and patients.The laboratory GC role in the precision medicine program was originally designated to answer high-level questions from patients and providers regarding the electronic consent, the enrollment process, and the genetic test. In addition, laboratory GCs help facilitate pre- and post-test genetic counseling related to the precision medicine program as needed. In the pilot phase, the laboratory GC contacted the ordering provider to discuss the MAP-positive results to help the provider discuss the information with their patient. The process was reassessed because some patients had not reviewed their MAP-positive result with their PCP prior to their clinical genetics consult. This led to the laboratory GC notifying both the provider and the patient of the results and placing a referral for all MAP-positive patients, as well as other genetic screen patients from the precision medicine program who had questions, to see a clinical GC or geneticist. The laboratory GCs replied to patients who requested a telephone response to their questions and acted as resources for the GCAs. They also worked closely with the laboratory directors on tasks such as a variant curation and interpretation, confirmatory testing logistics, and creating MAP-positive result reports.The precision medicine program motivated the expansion of clinical genetics into primary care. Clinical GCs were selected to be available to PCPs for informal consults, same-day patient appointments, and resources for anything genetic-related. The incorporation of a clinical GC into each region increased the genetics awareness and understanding among providers and patients. GCs also played a critical role in educating providers about genetic services, as described below.All patients who received MAP-positive results from the precision medicine program were referred to the clinical GCs or medical geneticists. Additionally, patients who had questions during enrollment, questions about uninformative results, or concerns regarding a personal or family history of a disease were referred.As the hub for patient care, it was critical to make PCPs aware of the precision medicine program and to empower them to respond to findings appropriately. PCPs were encouraged to engage with clinical GCs, who were embedded in primary care clinics, and were required to complete education modules designed to improve the clinician understanding of genetics . ProvideFrom its inception, the precision medicine program engaged in a multi-faceted approach to prepare all members of clinical teams for the introduction of genetic testing, from nurses to program directors. Efforts included educational sessions and clinic in-service presentations, which were provided by GCs and pharmacists early in the initiative, and later by GCAs. Each Sanford Health family medicine, internal medicine, and obstetrics and gynecology clinic was invited to three specific educational in-service presentations about the Imagenetics Initiative over the course of two years. Sanford Health also incorporated web-based training modules about genetics into the required annual and new-provider training for all physicians and advanced practice providers . Other eA novel aspect of Sanford Health’s efforts was a cooperative development of its future workforce. Sanford Health worked with Augustana University to create a genetic counseling master’s program, accredited in 2015, and a genetic counseling master’s program scholarship for students contracted to be employed at Sanford Health after graduation. This graduate program has provided genetic counseling students with an exposure to precision medicine through internships, thesis projects, and volunteer opportunities with the precision medicine program. Genetic counselors from the precision medicine program serve as program faculty and provide lectures related to the precision medicine initiatives. In addition, Sanford Health developed a pharmacy training program that exposes students and residents to pharmacogenetics. With two faculty member pharmacists on the PGx team serving as primary preceptors, the PGx team allows the opportunity for a select number of fourth year pharmacy students to do five-week clinical experiences as a PGx elective in either North Dakota or South Dakota. Post-Graduate Year 1 pharmacy residents at Sanford Health in Fargo, ND, and Sioux Falls, SD, are also offered the opportunity to complete a PGx elective block. In 2021, the PGx pharmacy team welcomed their first Post-Graduate Year 2 Clinical Pharmacogenomics Resident, a training position open to a new resident every year. This program aims to create a pool of pharmacists with PGx expertise and familiarity with Sanford Health that can continue to advance the mission of the precision medicine program.® Office Excel® 2016 . All active departments referring to a geneticist or genetic counselor from 01 January 2013 to 31 December 2020 were reviewed. Referrals made from pediatric departments were omitted from analyses because pediatric patients were not eligible for the precision medicine program. Referrals from genetic or laboratory departments were also omitted because these referrals reflected communication orders, such as referrals to the wrong department, rather than actual referrals. The departments that placed at least one referral order to a geneticist or genetic counselor were grouped into one of the following specialty areas: oncology, surgery, family medicine, internal medicine, obstetrics/gynecology, neurology, breast clinic, cardiology, endocrinology, otolaryngology (ENT), gastroenterology, nephrology, and other. The department specialty was determined by the name of the department. Some departments in rural areas with department names based on location rather than specialty were categorized as “other”.We used administrative records and EMR data to summarize the genetic referrals, uptake, and results of the precision medicine program. The data were summarized descriptively using MicrosoftThe protocol for the research summarized here is implemented in accordance with the Declaration of Helsinki and the ethical guidelines for medical research covering humans. The protocol was reviewed by the Sanford Institutional Review Board and determined not to be human subjects research (No. STUDY00002485/16 September 2021).The number of referrals to genetics and genetic counseling services at Sanford Health increased from 1438 in 2013 to 7517 in 2020 . The numAnticipating the growth of genetic services, an additional 1.0 full-time equivalent (FTE) clinical GC  was hireIn January 2013, clinical GCs primarily worked within maternal fetal medicine, oncology, pediatrics, neurology, and reproductive endocrinology. By December 2020, GCs had expanded into cardiology, internal medicine, and primary care settings. As the uptake of genetic screening increased, the lab GC roles added 3.0 FTE specifically for the precision medicine program to support the existing utilization management lab GC, accounting for a total of 3.6 FTEs . OverallMUTYH and ATP7B genes associated with autosomal recessive conditions, respectively.Of the 11,771 patients who enrolled in the precision medicine program through 31 December 2020, 11,505 (97.7%) agreed to receive screening for MAPs . One hunA medical geneticist and/or genetic counseling session was completed for 415 of 485 MAP-positive patients (85.6%). Thirty-three of 70 MAP-positive patients who did not meet with clinical genetics (47%) had prior genetic consults, 24 (34%) were non-responsive to attempts to schedule a genetic consult, and 9 (13%) canceled or did not come for a scheduled genetic consult. Four MAP-positive patients actively declined a genetic consult.Of the 11,680 patients from the precision medicine program with PGx data available through 31 December 2020, 11,095 (95.0%) had at least one atypical PGx finding. Of these patients, 1764 (15.9%) were identified by pharmacists to be on at least one medication for which there was an identified gene–drug interaction that warranted a further review by the prescribing provider. To handle the increased workflow for the precision medicine program, a Pharmacogenomics Manager and two clinical faculty members, each with 0.5 FTE devoted to the precision medicine program, were hired in 2018. In 2019, two full-time clinical pharmacists were added to the PGx team, and the manager position was transitioned into a director position overseeing the pharmacy, GC, and GCAs. In 2020, another 1 FTE PGx clinical pharmacist was added.Examples illustrating how team members collaborated to addressed results from the precision medicine program, primarily through monthly patient case conferences, are summarized in In this paper, we summarized the personnel changes that facilitated the integration of genetics into patient care across the Sanford Health system, including primary care. Sanford Health built a multi-disciplinary team that worked together to serve as a resource for PCPs and embraced clinical champions who helped to roll-out the Imagenetics Initiative. These personnel changes and proactive approaches to the workforce development, combined with provider education and EMR-based CDS, provided PCPs and other providers with the knowledge and support they needed to respond to the increased use of genetic testing. Importantly, the strategic plan Sanford Health enacted allowed the healthcare system to accommodate a large increase in genetic services with a modest increase in its genetic specialist workforce. Such plans are necessary given the expectations that shortages in the numbers of genetic specialists are likely to persist. Our results provide a unique insight into how a strategic development and the deployment of a multi-disciplinary clinical team during the implementation can empower a successful precision medicine program.Importantly, we observed a five-fold increase in referrals to clinical genetics compared to a three-fold increase in FTEs for clinical geneticists and genetic counselors from 2013 to 2020. We also observed increases in the number of individual departments providing referrals to genetic specialists. The increases in referrals may reflect the impact of educational efforts to make providers more aware of their access to genetics specialists and feel more prepared to act on genetic information. There are external factors that also likely contributed to the growth in referrals, including the improved capabilities of genetic tests , an incrOur data also demonstrate one model of how healthcare systems that pursue a greater integration of genetics into patient care should be prepared to adapt staffing over time. Initially, PCPs were expected to be the first providers to discuss results with their patients, but because this follow-up was not always occurring, laboratory GCs assumed responsibilities for the initial follow-up of MAP-positive findings. It is unclear if the lack of follow-up by PCPs was due to a discomfort about discussing results, if the PCPs think this is beyond their responsibilities, or if it is a lack of time given the many competing priorities of PCPs. Regardless, this finding highlights the importance of having genetic professionals as a resource for generalists and medical subspecialists outside of genetics.The findings also demonstrate the importance of provider education and workflows that allow genetic specialists to focus on patient care. Recognizing that genetics is a rapidly evolving area of medicine with known workforce shortages , SanfordAnother important lesson from the Sanford Health experience, highlighted by the vignettes, was the importance of a multidisciplinary approach to providing care and evaluating patient histories and genomic screening results. In many cases, different providers’ unique perspectives contributed to improved care, including cases where multiple-provider involvement was important for a single patient. A combination of genetic results, patient history, and critical thinking led to improved care. This work suggests that when healthcare systems implement a precision medicine program, hiring and supporting a variety of specialists is essential to optimizing patient care.While no single approach for the implementation of precision medicine programs is likely to be appropriate for all healthcare systems, our work highlights key points that should be addressed to increase the likelihood of success. Our experiences provide important lessons that other healthcare systems leaders can incorporate as they explore implementing their own clinical screening programs.


A preliminary comparison of the program experience and costs associated with the virtual interview season during the 2020-2021 COVID-19 pandemic against the traditional in-person interview process during the 2019-2020 interview season.Cross-sectional survey.Our institutional program launched an online survey via REDCap to otolaryngology programs across the country.A 33-item survey was sent to otolaryngology residency program directors regarding their experience and costs associated with virtual interviews during the 2020-2021 cycle and in-person interviews during the previous 2019-2020 cycle. Purchasing cost and opportunity cost were calculated for each program.P < .001) for a similar number of residency spots per program . The virtual interview led to more half-day interviews, a shorter duration of each interview, and fewer interviews completed per interview date. Purchasing cost decreased by $1940.46 (73%), and person-hours dedicated to the interview process decreased by 52.36 with the virtual interview. Total savings per program with virtual interviews were an estimated $6941.66.Twenty-two programs sent back completed survey responses. Program responses were equally represented among all regions of the United States. In the 2020-2021 interview season, programs received more applications (mean, 400 vs 336 the year prior, Virtual interviews in the setting of the COVID-19 pandemic led to a shift in application and interview patterns and was associated with a reduction in costs for programs when compared with the in-person interview format. In medical education, the entire otolaryngology residency interviewing season was conducted via virtual interviews, an unprecedented shift from the norm of the in-person interviewing format. Virtual interviewing offers advantages and disadvantages to applicants and residency programs. One significant advantage of virtual interviews for applicants is alleviation of the cost burden associated with in-person interviews, which includes the financial aspect and time involvement. Multiple studies have demonstrated how expensive the residency application process is for applicants, with financial costs in excess of $11,500.210The COVID-19 pandemic has transformed many aspects of otolaryngology and medical education. In the clinical arena, the deployment of telemedicine and virtual patient visits has skyrocketed.The in-person residency interviewing process poses significant costs to residency programs as well. Some of the associated costs include lost hospital revenue due to attending physicians’ inability to perform clinical duties on interviewing days, meal costs, and room and equipment rental. While the cost advantages of a virtual interview process from the applicant perspective have been well explored in the literature, there is a paucity of data analyzing the effects of virtual interviewing on residency programs. This study provides a preliminary review of the cost differences with the virtual interview format for otolaryngology residency programs by reviewing survey data collected from a sampling of residency programs throughout the country during the 2019-2020 and 2020-2021 interview cycles.We performed a cross-sectional survey-based study comparing the financial aspects and time involvement of the virtual interview format during the 2020-2021 COVID pandemic and the in-person interviews during the 2019-2020 cycle. This study was submitted to the University of California San Diego’s Institutional Review Board and approved for exemption status.Supplement 1, available online). The survey was modified from a previously published study that focused on the financial costs associated with the surgical residency interview process. In this previous study, surgical program directors were given a survey to assess program characteristics as well a variety of details related to conducting on-site interviews. The survey was verified and approved by the Association of Program Directors in Surgery’s Research Committee. This survey was then adapted for our study to gather data from the 2020-2021 and 2019-2020 cycles.A 33-item online survey was designed and sent to otolaryngology residency program directors requesting a comparison of their experience and costs associated with the virtual interview and the in-person interview format  was used for all analysis, and statistical significance was taken at P < .05.For the financial cost analysis, purchasing costs were defined as the costs of all goods purchased for interviews. Examples of purchasing costs include funds spent on meals, equipment rentals, and printing supplies, in addition to others. Opportunity cost was defined by costs associated with faculty being removed from clinical duty and other obligations to assist with interviewing of applicants. Opportunity cost was calculated in person-hours, or the number of persons involved in interviewing applicants multiplied by the number of hours worked. Average national wages for chairs, otolaryngology physicians, administrative staff, and residents were used in these calculations.A total of 22 programs completed all survey questions for a response rate of 21%. Program type and regions are presented in P < .001). There was a similar number of residency spots per program, with 3.04 available in the 2020-2021 cycle vs 3.0 in the 2019-2020 cycle . Despite similar numbers of invitations, programs completed more total interviews in 2020-2021 (47.23) than 2019-2020 . Fewer applicants canceled interviews after accepting the interview offer during the 2020-2021 interview cycle (5.23) as compared with the 2019-2020 cycle .For the 2020-2021 interview season, programs received a mean 400 applications as opposed to 336 for the year prior (P = .002). Programs decreased the number of interviews completed in an interview day from a mean 8.18 to 7.45 (P = .003). Additionally, each interview was shorter in duration at a mean 17.5 minutes in 2019-2020 vs 16.3 minutes in 2020-2021 . They spent significantly less on social events at a mean $1329.42 during 2019-2020 vs $137.50 during 2020-2021 (P < .001). They also spent significantly less on printing and supplies (P = .029) and shuttles (P = .039). There were no differences in purchasing costs for room reservations or venue and audiovisual costs (P < .001).Change in cost for programs with the virtual interview was analyzed. This was divided into 2 categories: purchasing costs and opportunity costs. For purchasing costs, programs spent a mean $1117.06 on food during the 2019-2020 interview cycle as opposed to $288.82 during the 2020-2021 interview cycle (al costs Figure 2Opportunity cost was then calculated in person-hours and average cost as described earlier Table 4.This preliminary study examines the effects of the virtual interview on various aspects of the interview process, including the numbers of applications, components of the interview, and costs associated with this format of residency interviews. It addresses certain methodological differences seen in the current literature and focuses on the program’s perspective rather than the applicant’s perspective with virtual interviews. As a cross-sectional study, the survey allows us to gather information from a sampling of residency programs to understand the changes to program costs associated with virtual interviews in the COVID-19 era.We found that the advent of the virtual interview led to more applications received per program with more total interviews completed. The virtual interview led to more half-day interviews, shorter duration of each interview, and fewer interviews completed per interview date. There were fewer interviews canceled by applicants. Programs spent less money on food, social sessions, printing, and shuttles, leading to almost $2000 in purchasing supply reductions during virtual interviews. In addition, fewer hours were spent by faculty and staff for the virtual interview process by a calculated average of 52 hours per program.31517 While the cost of interviews for residency programs has not been explored to the same extent as costs to applicants, program costs for residency interviews can reach upward of $100,000. Given these immense expenses, efforts to reduce the interview budget have been suggested. In 2010, the University of Arizona ophthalmology residency program conducted a prospective study where applicants were offered the choice to interview in person or virtually via video conferencing. These authors concluded that virtual interviewing is a viable, potentially cost-reducing option for applicants and programs. To reduce program costs further, they suggested conducting interviews in the evenings to limit lost clinical volume and revenue.Conversations related to the costs of residency applications and interviewing are certainly not new. It is well known that residency interviews are very costly for applicants, ranging from $2000 to $15,000 depending on the specialty and study conducted. While these data support that virtual interviews were well tolerated by applicants and faculty, multi-institutional studies that analyze the cost differences with virtual interviews for residency programs are lacking in our literature. Our study serves as an initial preliminary assessment of these cost differences for our residency programs.The COVID-19 pandemic created a unique situation where altering the interview format was no longer a conversation but rather a necessity. Residency and fellowship programs were forced to implement Zoom, Microsoft Teams, and other video conferencing technologies to conduct their interviews due to the public health crisis. A survey of surgical oncology fellows and faculty showed that 75% of applicants and 100% of faculty stated that the virtual interview process was “very seamless” or “seamless” and cited decreased cost, time saving, and increased efficiency as benefits to virtual interviewing. The travel and accommodation factors are a significant logistical and cost-based barrier otherwise. In addition, programs may have decided to increase the number of interviews to avoid unfilled residency spots as the effect of the virtual interview created much uncertainty. Despite more days of interviewing, the amount of costs on supplies decreased, while the time spent by the program director, associate program directors, faculty, and residents remained relatively unchanged. Costs associated with the virtual interview are not documented well in the literature, but our findings are consistent with Davis et al showing $500 to $1000 of cost savings with the virtual interview process.The popularization of the virtual interview due to the COVID-19 pandemic led to multiple changes shown in this study. The virtual interview format led applicants to apply to more programs and cancel fewer interviews. This shift correlates with the decreased time and cost barriers associated with the virtual interview as compared with the in-person format and is consistent with conclusions made by Siddiqui et al. Program directors will need to create strategies that will help them assess the fit of the candidates for their programs.The virtual interview led to a change in the interview components, with some programs canceling tours and social sessions and many no longer providing meals to applicants. This transition to a virtual format with less time to interact with faculty and residents in a more relaxed setting can affect a program’s ability to accurately assess a candidate as well as a candidate’s ability to assess a program. In a recently published study out of Wright State University, program directors felt that their ability to determine a candidate’s competitiveness in a virtual format was unchanged, but more subjective components, such as fit for their programs, were more challenging to assess during virtual interviews.23 Last, our sampling size limits us from having a more elaborate statistical analysis. However, our research provides an initial review of program costs that can help the conversations that are actively occurring in our specialty to ensure successful matches for applicants and programs in a cost-effective manner. A prospective study reviewing all otolaryngology programs’ experiences with the virtual interview format would provide us with the necessary guidance to navigate the changes to our interviewing process.There are multiple limitations associated with our preliminary study. First, survey results were sent to program directors after the interview season and could be subject to recall bias. Second, we focused our cost analysis strictly on measurable expenses, such as purchasing cost and opportunity cost. However, there are theoretical costs of virtual interviewing as well, such as a possible greater risk for a poor fit between a program and an applicant. This could lead to substantial increased costs for programs over time if the resident transfers out of the program due to a poor fit or disciplinary decisions. This cost is much more challenging to measure but may require programs to incorporate alternative interviewing strategies, such as a hands-on exercise or personality test.During the 2020-2021 virtual interview season, otolaryngology applicants applied to more programs and canceled fewer interviews. Residency programs conducted more interviews that were shorter in duration. Within our sampling of residency programs, virtual interviews created a mean financial cost reduction of $7000 per program, or 8.7% of their total interview costs. The opportunity cost of person-hours was associated with a reduction of $6942, or 13.8% decrease in mean total person-hours. Similar reductive trends in financial and opportunity cost are predicted with a larger study.Andrew Yousef, conceptualized the project in addition to leading the data collection and analysis components of the project and helped write the manuscript; Benjamin Bernard, assisted with literature review, data analysis, and data visualization and helped write the manuscript; Deborah Watson, conceptualized the project, oversaw project administration, assisted with data analysis and revisions of the manuscript.Competing interests: None.Sponsorships: None.Funding source: None.Click here for additional data file.Supplemental material, sj-pdf-1-opn-10.1177_2473974X221128908 for Virtual Interviewing in the Era of COVID-19: A Preliminary Analysis of Otolaryngology Residency Program Costs by Andrew Yousef, Benjamin Bernard and Deborah Watson in OTO Open: The Official Open Access Journal of the American Academy of Otolaryngology-Head and Neck Surgery Foundation


We found that the 5-HT7 receptor gene encodes two molecularly distinct transcripts, Msep5-HT7L and Msep5-HT7S, by the mechanism of alternative splicing in M. separata. Msep5-HT7S differs from Msep5-HT7L based on the deletion of 95 amino acids within the third intracellular loop. Two Msep5-HT7 receptor isoforms were activated by 5-HT and synthetic agonists α-methylserotonin, 8-hydroxy-DPAT, and 5-methoxytryptamine, resulting in increased intracellular cAMP levels in a dose-dependent manner, although these agonists showed much poorer potency and efficacy than 5-HT. The maximum efficacy of 5-HT compared to the two 5-HT isoforms was equivalent, but 5-HT exhibited 2.63-fold higher potency against the Msep5-HT7S than the Msep5-HT7L receptor. These two isoforms were also blocked by the non-selective antagonist methiothepin and the selective antagonists WAY-100635, ketanserin, SB-258719, and SB-269970. Moreover, two distinct mRNA transcripts were expressed preferentially in the brain and chemosensory organs of M. separata adults, as determined by qPCR assay. This study is the first comprehensive characterization of two splicing isoforms of 5-HT7 receptors in M. separata, and the first to demonstrate that alternative splicing is also the mechanism for producing multiple 5-HT7 isoforms in insects. Pharmacological and gene expression profiles offer important information that could facilitate further exploration of their function in the central nervous system and peripheral chemosensory organs, and may even contribute to the development of new selective pesticides.Serotonin  is an important neuroactive molecule, as neurotransmitters regulate various biological functions in vertebrates and invertebrates by binding and activating specific 5-HT receptors. The pharmacology and tissue distribution of 5-HT receptors have been investigated in several model insects, and these receptors are recognized as potential insecticide targets. However, little is known about the pharmacological characterization of the 5-HT receptors in important agricultural pests. In this study, we investigated the sequence, pharmacology, and tissue distribution of  Drosophila melanogaster Meigen (Diptera: Drosophilidae) and Gryllus bimaculatus De Geer (Orthoptera: Gryllidae) [D. melanogaster and Apis mellifera Linnaeus (Hymenoptera: Apidae) [D. melanogaster and Manduca sexta (Linnaeus) (Lepidoptera: Sphingidae) [Serotonin  is an important biogenic amine, as the neurotransmitter/neuromodulator participates in various critical physiological functions in both vertebrates and invertebrates ,2,3,4. Fyllidae) ,7,8; fee Apidae) ,11,12,13ingidae) ,15,16. i/o proteins, and thus inhibit cyclic adenosine monophosphate (cAMP) synthesis [q/11 proteins, causing an increase in cytosolic Ca2+ concentration [s proteins, activating adenylyl cyclase (AC) activity and hence increasing intracellular cAMP formation [Pieris rapae (Linnaeus) (Lepidoptera: Pieridae) that mediated increased intracellular calcium concentration, which may be an invertebrate-specific receptor without a counterpart in mammals. The pharmacological characteristics of the same type of 5-HT receptors vary significantly among different insect species [Haemonchus contortus (Rudolphi) Cobb (Rhabditida: Trichostrongylidae) could inhibit the growth or kill the larvae of Mythimna separata  (Lepidoptera: Noctuidae).In vertebrates, the effects of serotonin are mediated by its binding to two specific types of receptors: ionotropic and metabotropic receptors. All metabotropic receptors belong to G protein-coupled receptors (GPCRs), with seven transmembrane domains that are divided into six distinct classes based on amino acid sequence homology, pharmacological properties, and downstream second messenger pathways , while the remaining 5-HT3 receptor is an ionotropic receptor ,18. Withynthesis . The 5-Hntration ,19, whilormation  recogniz species ; therefo species  demonstr5-HT7 receptor genes have been cloned in many vertebrates. In humans, mice, and rats, the coding region of this gene contains three introns, and alternative splicing at the second and third introns is a general mechanism that results in a number of splice variants. No major differences in their pharmacology or functionality have been reported so far [s proteins and stimulate cAMP formation [5-HT7 receptor gene transcript has been cloned in insects thus far [D. melanogaster [A. mellifera [Calliphora vicina Robineau–Desvoidy  [The d so far , and allormation . On the thus far . Tissue thus far ,24,25. Tnogaster , learninellifera , and salhoridae) . The pha5-HT7 receptor gene from M. separata, one of the important polyphagous insect pests in China, costing more than ¥ 146 million ($18 million) in insecticide management on wheat alone in 1997 [5-HT7 receptors gene transcription patterns in other insects [M. separata. To our knowledge, this is the first report showing that alternative splicing is the mechanism for producing multiple 5-HT7 isoforms in insects. Furthermore, we examined the pharmacological characteristics and expression profiles in different tissues of M. separata larvae and adults. These results provide a foundation for understanding the actions of serotonin in behavior and physiological function, and for exploring novel pesticides to control this pest in the future.In the present study, we cloned the  in 1997 . Differe insects ,27,28, tMsep5-HT7 from adult M. separata was amplified by using transcriptome analysis with a PCR-based strategy. The Msep5-HT7 gene contains an intron and produces two molecularly distinct transcripts (Msep5-HT7L and Msep5-HT7S) by the mechanism of alternative splicing  (Lepidoptera: Bombycidae), and 77.6% for Papilio Xuthus Linnaeus (Lepidoptera: Papilionidae). The similarity between Msep5-HT7S and lepidopteran insect 5-HT7 orthologues is 78.5% for M. sexta, 76.8% for P. rapae, 77.0% for B. mori, and 64.3% for P. xuthus. This clearly shows two Msep5-HT7 receptor isoforms clustered with other insect 5-HT7 receptors in a distinct clade, indicating that they belong to the 5-HT7 type receptors by phylogenetic tree analysis  of cAMP production was achieved at 5-HT concentrations of 9.24 × 10−8 M (log EC50 = −7.03 ± 0.10) and 3.51 × 10−8 M (log EC50 = −7.46 ± 0.19) in Msep5-HT7L-expressing and Msep5-HT7S-expressing cells, respectively  and synthetic 5-HT receptor agonists  at a final concentration of 10 μM to alter intracellular cAMP levels in HEK293 cell lines that stably express either Msep5-HT7L or Msep5-HT7S receptors. Among the four tested biogenic amines, only 5-HT significantly increased cAMP levels in cells expressing either Msep5-HT7L or Msep5-HT7S receptors. All synthetic agonists activated the Msep5-HT7L receptor, while only αm-5-HT significantly increased cAMP levels in cells expressing the Msep5-HT7S receptor, but showed lower efficacy than 5-HT A,B. We eectively . 50 values of the synthetic agonists were more than 100-fold higher than those of 5-HT, implying that they were considerably less potent than 5-HT  of synthetic agonists were also consistently poorer than the same concentration of 5-HT induced responses, and the efficacy of synthetic agonists was different between the two receptor isoforms. The highest concentration of αm-5-HT and 5-MT (1 mM) resulted in about 69% activation compared with the maximum amount of cAMP stimulation obtained with the Msep5-HT7L receptor with the same concentration of 5-HT. However, these two agonists resulted in about 35% activation of the maximum amount of cAMP obtained in the Msep5-HT7S receptor. The highest efficacy of 8-OH-DPAT was below 30% of the maximum response of 5-HT in the two isoforms C,D. All han 5-HT . The agohan 5-HT .50) values are listed in Potential synthetic antagonists were tested by simultaneously applying 5 μM of 5-HT and an 8-fold dose of antagonists to stable transfected cell lines expressing Msep5-HT7L or Msep5-HT7L receptors. All of the tested mammalian 5-HT receptor antagonists  decreased the 5-HT induced response in Msep5-HT7-expressing cell lines by about 33–60% A,B. The Msep5-HT7 gene has two distinct transcripts, Msep5-HT7L and Msep5-HT7S, which are generated by alternative splicing. The specific primers that amplify the Msep5-HT7 gene contain these two transcripts (Msep5-HT7S and Msep5-HT7L in different tissues of larvae and adults were quantified by qPCR (relative expression level of Msep5-HT7S = relative expression level of Msep5-HT7 − relative expression level of Msep5-HT7L). The two transcripts were co-expressed in all tested tissues   .5-HT7 gene in M. separata that codes two molecularly distinct transcripts of 5-HT7 by the mechanism of alternative splicing, complying with the typical GU-AG rule. Msep5-HT7S differs from Msep5-HT7L based on an extra 3′-terminal and 285 nucleotides cleaved off during autocatalytic excision of the intron. The alternative splicing forms of 5-HT7 receptors are widely found in humans, mice, and rats, and alternative splicing occurs extensively at the second and third of three introns of the 5-HT7 gene [5-HT7 receptor genes have been cloned and characterized from D. melanogaster [Aedes aegypti (Linnaeus) (Diptera: Culicidae) [A. mellifera [C. vicina [M. sexta [Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae) [P. rapae [Plutella xylostella (Linnaeus) (Lepidoptera: Plutellidae) [5-HT7 genomic sequence in different insect species. We checked the coding region of the 5-HT7 gene from B. mori, T. castaneum, M. sexta, P. rapae, P. xylostella, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae), and Trichoplusia ni (Hübner) (Lepidoptera: Noctuidae) according to the genomic and coding sequence data. Only one intron was found in these insect 5-HT7 genes, which was located in the sequence encoding the putative third intracellular loop. We need to check whether this kind of alternative splicing is a common mechanism in insect 5-HT7 genes in the future. Two Msep5-HT7 receptor isoforms also contain key sequence motifs that are important for ligand binding and signal transduction for biogenic amine receptors, compared with previous reports [We cloned one HT7 gene ,31,32. Mnogaster , Aedes alicidae) , A. mellellifera , C. viciM. sexta , Triboliionidae) , P. rapaP. rapae , and Pluellidae) . Only on reports ,40,41,42M. separata, indicating that these receptor isoforms might have similar signaling pathways. The pharmacological properties of the two receptor isoforms were further explored by testing their downstream signal responses to synthetic agonists and antagonists. The efficacy of 5-HT on the two M. separata 5HT7 receptor isoforms was equal, but the potency of 5-HT was nearly three-fold higher against the Msep5-HT7S receptor than the Msep5-HT7L receptor. A similar potency of 5-HT against the 5-HT7 receptor has also been observed in other insects, such as M. sexta (EC50 = 10.5 nM), P. rapae (EC50 = 15 nM), T. castaneum (EC50 = 27.3 nM), and D. melanogaster (EC50 = 60 nM) [M. separata 5-HT7 receptor isoforms. The 5-HT7 receptor in animals is known to stimulate AC activity, which converts ATP to cAMP, and its coupling with specific G-proteins is brought about by amino acids in close proximity to the plasma membrane of the second and third intracellular loops and the cytoplasmic tail of the receptor proteins ,27. This= 60 nM) ,35,36. TT. castaneum [Our results are consistent with previous pharmacological studies on 5-HT7 receptors in other insects ,32,33,43astaneum . All thrM. separata 5-HT7 receptor, but their inhibitory potential against 5-HT7 receptors from different insect species were quite different. Methiothepin, a non-selective antagonist against mammalian 5-HT and dopamine receptors, showed about 70- and 480-fold higher inhibitory potency against 5-HT7 receptor from A. mellifera (IC50 = 0.13 μM) and P. rapae (IC50 = 0.90 μM), and 125-fold lower inhibitory potency against 5-HT7 receptors from T. castaneum (IC50 = 7910 μM) than Msep5-HT7S receptors (IC50 = 62.86 μM) [D. melanogaster, and C. vicina 5-HT2A receptors [Caenorhabditis elegans Maupas (Rhabditida: Rhabditidae), C. vicina, and T. castaneum, and it showed about 20- to 50-fold higher inhibitory potency against the 5-HT7 receptor from C. vicina (IC50 = 1.50 μM) and T. castaneum (IC50 = 1.95 μM) than to Msep5-HT7L (IC50 = 46.91 μM) and Msep5-HT7S (IC50 = 84.94 μM) [Periplaneta americana (Linnaeus) (Blattaria: Blattidae) [M. sexta, A. mellifera, and T. castaneum [T. castaneum (IC50 = 76.8 μM) [A. mellifera and C. vicina, and moderate inhibitory potency against the 5-HT1 receptor from T. castaneum (IC50 = 205 μM) [C. vicina (IC50 = 9 nM), P. rapae (IC50 = 1.04 μM), and T. castaneum (IC50 = 15.82 μM) [Five selective or non-selective antagonists of mammalian biogenic amine receptors were effective against 2.86 μM) ,36. Ketaeceptors ,45. It i4.94 μM) ,36,46. Wattidae) , and it astaneum ,35,39. I76.8 μM) ,50. SB-2 205 μM) ,27. Howe5.82 μM) ,36. Few studies have investigated the effects of SB-258719 on insect 5-HT receptors to date. The antagonistic potency of these two selective 5-HT7 receptor antagonists was statistically equal in terms of the half maximal inhibitory concentration against Msep5-HT7L and Msep5-HT7S. In our study, when the concentration of five antagonists increased 128-fold over 5-HT, they completely blocked 5-HT7 receptor-mediated cAMP production. These antagonists could be used as candidate agents for functional studies. The differences in the potency and efficacy of agonists and antagonists against the 5-HT7 receptor between mammals and insects, and even between different insects, imply that these types of GPCRs could act as potential targets for the development of selective insecticides. 5-HT7 receptor genes were detected in the Malpighian tube, gut, salivary gland, brain, and/or antennae of A. mellifera, G. bimaculatus, M. sexta, A. aegypti, C. vicina, T. castaneum, and Polyrhachis vicina Roger (Hymenoptera: Formicidae) [5-HT7 receptor genes are highly expressed in larval Malpighian tubes and guts, indicating that the 5-HT7 receptor potentially has a role in digestion and metabolism. Yang et al. [5-HT7 receptor’s mRNA and protein expression were slightly higher in the pupal than the egg and larval stages, and they deduced that this receptor may be involved in modulating adult formation and caste differentiation in P. vicina. Moreover, the 5-HT7 gene showed strong expression in the brains of adult D. melanogaster, and it mediated normal courtship and mating behaviors [5-HT7 were found in adult males than females of P. vicina and P. xylostella, as well as in our detected male M. separata antennae and labial palps. In addition, 5-HT-immunoreactive neurons were found to be distributed in the antennal lobe of M. sexta. It was then confirmed that 5-HT changes K+ conductance in the cells and increases the excitability and responsiveness of central olfactory neurons to sex pheromones [5-HT7 receptor genes in male antennae and labial palps should receive careful consideration, as it may contribute to modulating sensitivity and specificity to carbon dioxide and plant volatiles, perception of sex pheromones, and regulating courtship and mating behaviors. High expression of Msep5-HT7S in male antennae and labial palps could be attributed to its higher sensitivity to endogenous 5-HT than Msep5-HT7L receptor, and organisms that express Msep5-HT7S transcript can improve energy efficiency. High transcription levels of micidae) ,36,51,52ehaviors . We alsoeromones ,53, althM. separata is a typical seasonal, long-distance migratory polyphagous pest that can attack nearly 100 families of more than 300 kinds of food and industrial crops, resulting in serious yield losses in China and other parts of Asia and Oceania [The oriental armyworm  Oceania ,55,56. T Oceania . Thus, t Oceania ,22. WithM. separata were reared on an artificial diet in the climate incubator and maintained under a photoperiod of 14:10 h (light:dark) at 27 °C and 75% relative humidity [The larvae of humidity . Adult mM. separata using an EasyPure Genomic DNA Kit  and TRIzol reagent , respectively. Genomic DNA and single-stranded cDNA synthesized from the RNA using the Fast King gDNA Dispelling RT SuperMix  were used as the template for PCRs to amplify the genomic DNA sequences and open reading frame (ORF) of Msep5-HT7, respectively. Full-length sequences of putative Msep5-HT7 were obtained from our previous brain transcriptomes of M. separata [Msep5-HT7L (GenBank Acc. OM025087) and Msep5-HT7S (GenBank Acc. OM025088).Genomic DNA and total RNA were extracted from adult separata . The genM. separata 5-HT7 receptor, the phylogenetic trees of 5-HT receptors from other insects were constructed using MEGA 6.0 software. The maximum-likelihood method was used with a bootstrap analysis of 1000 replicates, based on the JTT matrix-based model [Drosophila melanogaster FMRF amide receptor was used as outgroup. The amino acid sequences used in the phylogenetic analyses are listed in Alignments of 5-HT7 receptor amino acid sequences were performed by MAFFT online, and the results were displayed by ESPRIPT . The traed model . The DroMsep5-HT7L and Msep5-HT7S cDNA containing the Kozak consensus sequence (GCCACC) [KpnI and XhoI , and after purification, they were cloned into a pcDNA3.0 vector . DNA sequencing was used to confirm the correct recombinant plasmid.The construction of expression plasmids of the (GCCACC)  were obt2. Four microgram recombinant plasmids with 10 μL of lipofectamine 2000  were introduced into exponentially growing HEK 293 cells. The 0.8 mg/mL antibiotic G418  was added to the medium to select for cells that were stably expressed the Msep5-HT7 receptors. After fourteen days of G418 selection, three G418-resistant colonies were trypsinized in cloning cylinders and transferred to 12-well plastic plates for expansion [HEK-293 cells were cultured in D-MEM  supplemented with 10% FBS  at 37 °C and 5% CO6 cells/mL, culture media were removed and the cells were washed two times with cold PBS (pH 7.4), then the cells were preincubated in 450 μL of Dulbecco’s phosphate-buffered solution (DPBS) containing 100 μM phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine (IBMX) for 20 min at room temperature. After the preincubation, 50 μL of DPBS, containing various concentrations of reagents, was added into the stable transfected cells for incubation for 30 min at 37 °C. The cells were washed three times with cold PBS (pH 7.4) and then lysed in 250 μL of cold cell lysis buffer by re-freezing. The cell debris was removed by centrifugation at 600× g for 10 min at 4 °C. The amount of intracellular cAMP extracted from harvested supernatant was determined using an ELISA reagent . Each measurement was performed in duplicate, and three independent assays were carried out for each reagent and concentration tested.Intracellular cAMP concentrations were measured as previously described ,63,64. BMsep5-HT7 gene in larvae and various adult tissues. cDNA was synthesized using total RNA that was extracted from above collected samples with the FastKing gDNA Dispelling RT SuperMix . The specific qPCR primers used to amplify Msep5-HT7 and the reference genes are listed in β-actin (GenBank Acc. GQ856238.1) and glyceraldehyde-3-phosphatedehydrogenase (gapdh) (GenBank Acc. HM055756.1) were used to normalize Msep5-HT7 gene expression. Reactions for each sample (20 μL) consisted of 10 μL of 2 × SuperReal PreMix Plus , 7.6 μL of sterilized H2O, 0.5 μL of each primer (10 μM), 0.4 μL of Rox reference dye, and 1 μL of cDNA. qPCR was carried out with an initial denaturation at 95 °C for 3 min, followed by 40 cycles at 95 °C for 15 s and 60 °C for 30 s. The qPCR reaction of each sample was performed in three technical replicates and three biological replicates. Then, we used a relative quantitation method (2−ΔΔCT) [Real-time PCR was used to confirm the expression profiles of the (2−ΔΔCT)  to evalup < 0.05) to analyze the expression differences of the Msep5-HT7 gene among different larvae and adult tissues. Then, we used Student’s t-test to test for statistically significant differences in Msep5-HT7 expression levels and expression ratio of 5-HT7S between the same tissues from male and female adults, respectively (p < 0.05).GraphPad Prism 7.0 software was used for curve fitting and statistical analysis. We used one-way analysis of variance (ANOVA) with Tukey’s multiple comparison test (


In the fall of 2013, the authors received funding to help develop and implement an afterschool wellness program alongside Indigenous youth aged 6–10 years old in the North Central neighborhood of Regina, Saskatchewan, Canada. The Growing Young Movers (GYM) afterschool program was funded, in part, as a corrective response to a broader social trend in which Indigenous youth in this neighborhood reported declining health and wellness outcomes, as well as multiple other barriers to social inclusion. This article discusses the reflections of three senior high school Indigenous youth (16–18 years old) who participated in the afterschool program as peer-mentors over a 2-year period from 2015 to 2017. Our inquiry reveals how these youth viewed the program—and their role(s) within it—in far more complex, active, and even political terms, than the program's initial framing as a physical activity-based “intervention” had anticipated. Our analysis (re)positions youth according to their own personalized voice and narratives as: cultural leaders, knowledge holders, and as agents of change in their community. Education scholars have drawn important attention to the merits of a “wrap-around” approach in afterschool youth programs to balance schoolwork, career training, and employment opportunities, with various other social activities, such as sports and other extracurricular activities Kugler, . EducatiIn the fall of 2013,This paper discusses the experiences of three high school Indigenous youth who all served as peer-mentors within the afterschool program over a 2-year period from 2015 to 2017. Adopting narrative inquiry as our primary methodology, our study revealed that the three Indigenous peer-mentors defined their experiences according to a series of deliberate and largely empowering agendas in which they envisioned themselves as cultural leaders, knowledge holders, and as active agents of change within their community. These findings stand in contrast to dominant narratives that often persist within popular and some scholarly accounts of afterschool programming in which youth participants are cast as passive consumers of a program's official messaging , with Indigenous youth under 20 years old representing nearly half of Regina's Indigenous population  provided the core content areas around which the afterschool program was structured. Community Elders also voiced the need for programming to enhance students' cultural wellbeing through purposeful movement opportunities connected to traditional land-based activities. The feedback from this collaborative process further resulted in an inter-generational approach to mentorship within the program—an approach that engaged Indigenous children (primary elementary students), Indigenous youth (middle and high school students), volunteers (university pre-service teachers), as well as the researchers, in various socio-cultural and physical activity experiences within the gymnasium and broader community.Each Wednesday for ~2-h, at the end of the school day, a group of 20–25 elementary-aged Indigenous youth and 6 senior high school peer-mentors met at the school gymnasium to engage in physical activity led by Brian (the first author). Brian co-facilitated these weekly sessions alongside the youth mentors, who also helped to organize snacks, group outings and discussions, and various other social and cultural activities. While the school gymnasium provided the primary meeting spot for our weekly activities, the group also traveled to various other locations throughout the city . Some of our more common activities included developmental games and physical activities that were modified to optimize student engagement across all ages and skill levels. The group also regularly engaged in guided conversations led by the peer mentors that prioritized youth voices to ensure that all activities remained inclusive, and youth driven.In terms of qualitative data, the research team accumulated a variety of field texts over the 4-year period, including observational data, conversational data, digital stories, and program evaluation surveys. In this paper, we narratively inquire into the conversational data to shed light on the experiences of three Indigenous youth mentors from the afterschool program: Candice, Clary, and Colinsocial (what was happening around them), temporality (how their experiences were bound in time), and finally, place, “which attends to the specific concrete physical and topological boundaries of inquiry landscapes” (p. 51). Brian framed the research puzzle with “a sense of a search, a ‘re-search,' a searching again… a sense of continual reformulation” constructed their identities. Following Clandinin These three lines are part of a short piece that Clary wrote as an introduction to be read to the children in the afterschool program a few months after she herself had joined. Clary and Brian revisited these words whilst reviewing her narrative account one final time. Curiously, Clary elected to rearrange the words to reflect a popular stereotype of Indigenous youths in North Central Regina as hopeless and flawed:isn'talways easypossible.Greatness Do notDoubt yourself.Anyone can be successful so long as you want to be are not Indigenous.Clary's edited version of the original text revealed her frustration with how Indigenous youths in North Central have been stereotyped. Colin expressed similar frustrations with how Indigenous youths have been treated in the area: “…sometimes when I'm in a store or something, there'd be people following me around. I don't really like it very much” Candice grew up hearing these narratives and referred to racial profiling as misguided assumptions that unfairly position Indigenous youth as risks to both themselves and to the broader public: “Many people assume what First Nation people are... they didn't finish school, they don't have jobs, they're living off welfare and they have lots of kids and maybe alcohol and drugs” . She sensed the intrusive and judgmental gaze of non-Indigenous neighbors each time the GYM program ventured beyond the North Central community:When we go out, when we take the kids on outings, it's like mostly when people see us, it's like a bunch of First Nations kids and First Nations mentors, and they assume that it's going to be hard with us there, or they're going to have troubles with us.For Brian, the racialized stereotypes confronted by Candice, Clary, Colin, contrasted markedly with the roles and identities exhibited within the afterschool program. Brian knew Colin as a leader—a person he could depend on and to whom the kids looked up and literally clung to. Candice, likewise, exhibited so much compassion for her younger siblings and proved to be a natural mentor for many of the other kids in the GYM program. Yet, it was clear that Colin, Clary, and Candice were negatively impacted by the racism and stereotypes they encountered on a daily basis. As Clary explained,Would you think that I was going to be successful in high school? Probably not. Because if you tell people my story, there's a lot of key bullet points that would say that I shouldn't be successful .retold and relived stories according to their own personalized narratives that further exhibited their leadership, compassion, and resiliency.These bullet points are emblematic of a deficit narrative that all three youths described as a perpetual obstacle that impeded their social advancement, and that also impeded their mental health and wellbeing. Brian's conversing with the youth through the weekly afterschool program helped him to understand the multiple landscapes upon with these deficit narratives impacted youths. However, these youths also The youths gradually felt comfortable sharing their stories with Brian once they had developed strong relationships through regular attendance at the weekly afterschool program. Brian noticed that the experiences lived and shared in the program were largely connected to feelings of belonging and fellowship that stood in contrast to the experiences of competition, winning, and losing that predominates many other youth sport programs. Instead of competition and aggression, the youths emphasized mentorship and role-modeling as key drivers of their involvement in the program, and as significant departures from the grand narrative associated with North Central Regina. For example, Clary explained how the afterschool program exhibited for youths an alternative narrative of education and career-development:Yeah, I think it gives them something to look … like role model, like someone to look up to whether they're just doing a one-on-one situation or playing with them all. I think, sometimes, especially a lot of them that are First Nation kids, when I was a kid, you don't meet a lot of First Nation people that are successful in their high up careers like being a doctor and stuff like that. I think if they see high school students working toward going to university, and university students working toward their career goal, then they'll realize that that's okay and that's possible and whatnot. That's ok and that's possible were words that resonated with the teachings of Joseph, an Elder affiliated with the afterschool program, who explained that “youths live the story they are told.” Clary's story centered on perseverance, resilience, and a desire to make a positive impression on the world. This was a story that was both shared and modeled by her mother—a role-model whom she tries to emulate through living her own mentorship story. As Clandinin . Maybe it doesn't matter if you're the slowest one in the gym, you'll always be included. We all get a chance to be strong if we work together .For Colin, the lived and told stories of North Central Regina must be rewritten to promote inclusivity, not division. Colin redirected his energy and commitment to youth and community development to the afterschool program. For Colin, Candice, and Clary, the afterschool program afforded a prime opportunity to both express and impart the behaviors they valued, thereby contributing to rewriting the narrative associated with North Central. As Clary described:I want the GYM program to instill a sense of belonging, because it's more like a family or sense of community. Because that's how community is, right? That's how family is, right? Everyone's not always the same age in a family. There's lots of differences and different things going on in a community, but we're still connected. That's how I want everyone to feel when they come to GYM, that they belong here. Clary's reflection reminded Brian of the stories she had shared about her childhood. As noted above, Clary had a great deal of reverence for her mother, a single parent, who treated everybody equally and who supported the family through thick and thin. Clary modeled her relationship with others in the program in a similar fashion: “Supporting each other, that's what families do, right!” (Narrative account negotiation). Clary, along with the other mentors, not only shared this narrative with youth in the afterschool program, they lived it every day.For example, Candice prided herself on being a “good” big sister who takes care of her siblings. She embraced a similar guardianship role in the afterschool program, and prided herself on making sure everyone felt safe and included in the various activities. Candice also explained how caring for others was not a burden, but a privilege that afforded her great joy:I enjoy seeing kids have fun and play, so it kind of makes me feel happy. It makes me feel like if I play with them, it makes me feel young, I guess. I don't know. I like seeing little kids happy, so it makes me happy. Candice further explained how she often felt lonely as a child, and identified the afterschool program as an opportunity to help prevent that feeling of loneliness in others: “It's important because we want all the kids to feel happy and feel like they belong. We make sure that everyone feels like they're being included in the activities. I know how it feels to not be included.”  The undercurrent of Candice's ‘stories to live by' is rooted in a strong sense of guardianship, community, and a deep compassion for others. Her narrative prioritized nurturing a spirit of belonging among youths from diverse backgrounds and domestic situations—a teaching she credited to her Aunt, Ruth.As noted above, Candice dedicated herself to ensuring that all youths felt safe and valued in the afterschool program. Brian learnt that, from an early age, Candice had lived with her Aunt Ruth, who was in fact her foster parent. Affectionately known to all her foster children as “Auntie”, Aunt Ruth proved to be a tremendous role model and source of support for Candice and her foster siblings from the age of 8 years old. According to Candice, Ruth was caring and compassionate, and treated all her children equally. In addition, Ruth also taught Candice important life lessons about how to behave and act in public:I think she taught me a lot about how to be respectful and an overall better person. She taught me how to be proper … like, teaching me how to have manners and that kind of thing. She also taught us all how to act in public, stuff like that. (1. p. 11)Candice wanted to pass on Ruth's teachings to the other youths in the afterschool program, especially those who may not have enjoyed the same quality of mentorship that she had experienced as a youngster. For Candice, the afterschool program was about preparation for life beyond the gymnasium. In that sense, she prided herself on promoting teachings that went beyond physical activity within the afterschool program:I want them to grow as individuals. That's important. Remember, we're also teaching them how to be polite, how to have manners, how to be respectful. School isn't really focusing on basic stuff like that, and that's kind of what we're doing so they learn those manners in the program and can better succeed in and outside of the school. Of significance in the above excerpt is that Candice positioned herself as a teacher and as part of a larger teaching community accountable to those youths who showed up each week to the program. Candice also recognized Aunt Ruth as the source of her knowledge and she sought to honor her teachings through passing them on to others in the program.Colin exhibited a similar commitment to teaching youths lessons that would help prepare them for life—lessons that he also believed were insufficiently addressed in school:Well, in school, teachers usually teach what they think you should be learning. Education is what you should be learning, right? There's a difference. What they teach and what you will need in your future may be two entirely different things. Anyone can teach you math, like 1 + 1, but can math really teach you generosity? Those sorts of lessons are shown to you by family, friends, and community, right? Not in a classroom real important stuff in life. That's the stuff that can take you places” .For Colin, the lessons that were typically preached in school did not always square with the lessons required for life beyond the classroom environment. Education involved learning “real life stuff” . “It's part of our life and begins the day we are born.” . Like Candice, Colin credited a family member—Grandma Jean—with teaching him the most important lessons about love, compassion, and self-respect. “My grandma, she really helped me understand everything, like how to look out for others and treat people with respect. That's the For both Candice and Colin, education unfolded across multiple landscapes; e.g., institutional, familial, and community landscapes. Colin shared an experience from his elementary school physical education class, which, incidentally, took place in the same gymnasium that hosts the afterschool program. Brian asked if the gymnasium felt any different from when Colin had attended the school as a youngster. Colin explained, “Well, there's only one teacher in a physical education class. But now, in our program, we've got five, six, seven teachers sometimes. Even the kids play a big role in teaching other kids. I think our program approaches teaching and learning as a community, not as an individual. That's a big difference. We also draw our teachings from everywhere, right? We're not reading a government curriculum. Our teachings come from us” . When asked if he considered himself to be one of those teachers, Colin replied with a big smile across his face, “yeah, for sure.” He further explained,A lot of these kids, they don't have older brothers. Or, if they do, they don't really interact with them. Everyone needs to look up to someone or something. When we're playing with these kids, we're helping them. By doing that, it puts us in the role of that older brother or older sister. I think that a lot of the younger kids look up to us older kids. They see us as teachers and Elders, or maybe as an older brother or sister. We have a responsibility to bring them along as best we can by sharing our best teachings. Colin, Candice, and Clary all felt a great deal of pride and responsibility for mentoring youths in the afterschool program. They envisioned themselves as teachers, Elders, older siblings, and as part of a broader teaching and learning community invested in sharing their wisdom with the next generation of youths—an investment that they firmly believed would lead to positive social change both in and beyond their own lives.In this article, we have tried to position Indigenous youths not as passive consumers of dominant messaging, but as co-authors of the afterschool program. In so doing, we have shed light on an area of youth development that remains underexplored; that is, how youths experienced an afterschool program intended to enhance their overall health and wellness (see Bruner et al., As we have written about previously, (Schaefer et al., The original contributions presented in the study are included in the article/supplementary materials, further inquiries can be directed to the corresponding author/s.The studies involving human participants were reviewed and approved by University of Regina, REB. Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin.LS was BL's Ph.D. supervisor while this research was conducted. SL contributed to this work, as well as earlier drafts of this paper. JK was added as an author at a later date, and contributed quite significantly to this final draft. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.