Brazil is home to the highest absolute number of human T-cell lymphotropic virus type-1 (HTLV-1)-infected individuals worldwide; the city of Salvador, Bahia, has the highest prevalence of HTLV-1 infection in Brazil. Due to the complex nature of several diseases associated with this retrovirus, a multidisciplinary health care approach is necessary to care for people living with HTLV-1. The Bahia School of Medicine and Public Health’s Integrative Multidisciplinary HTLV Center (CHTLV) has been providing support to people living with HTLV and their families since 2002, striving to ensure physical and mental well-being by addressing biopsychosocial aspects, providing clinical care and follow-up, including to pregnant/postpartum women, as well as comprehensive laboratory diagnostics, psychological therapy, and counseling to family members. To date, CHTLV has served a total of 2,169 HTLV-infected patients. The average patient age is 49.8 (SD 15.9) years, 70.3% are female, most are considered low-income and have low levels of education. The majority (98.9%) are HTLV-1 cases, and approximately 10% have been diagnosed with tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM), while 2.2% have infective dermatitis and 1.1% have adult T-cell lymphoma. In all, 178 pregnant/postpartum women [mean age: 32.7 (±6.5) years] have received care at CHTLV. Regarding vertical transmission, 53% of breastfed infants screened for HTLV tested positive in their second year of life, nearly 18 times the rate found in non-breastfed infants. This article documents 20 years of experience in implementing an integrative and multidisciplinary care center for people living with HTLV in Bahia, Brazil. Still, significant challenges remain regarding infection control, and HTLV-infected individuals continue to struggle with the obtainment of equitable and efficient healthcare. Human retroviruses were identified about four decades ago. Human T-cell lymphotropic virus type-1 (HTLV-1), identified four years before the successful isolation of human immunodeficiency virus (HIV), was the first human retrovirus associated with the clinical development of disease . HTLV-2 Efforts by Brazilian scientists have contributed to a better understanding of the epidemiology, clinical and laboratory aspects as well as pathogenesis of this infection, thereby raising awareness of HTLV-1 as a serious public health problem in Brazil . AlthougIt is important to note that while there is no cure or effective vaccine for HTLV-1 infection, pharmacological and non-pharmacological treatments help minimize patient suffering and improve the quality of life (QoL) of this neglected population. Furthermore, while additional public health measures are required to prevent and/or control the spread of HTLV-1 infection, health care provisions for patients also remain a challenge , 23.Human T-cell lymphotropic virus type-1 infection can cause proliferative disease, such as adult T-cell leukemia, and inflammatory disorders, including HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), uveitis, and infectious dermatitis –28. OthePeople living with human T-cell lymphotropic virus may also present a higher prevalence of psychiatric/psychological disorders than the general population, including signs of psychological distress, anxiety, sleep and psychosomatic disorders, suicide ideation, low self-esteem, and depression –43. IndeThe Bahiana School of Medicine and Public Health (EBMSP), created in 1952, is a private, non-profit higher education institution dedicated to teaching, research and providing health-based extension programs. The EBMSP curriculum consists of seven undergraduate courses . EBMSP also develops scientific, cultural, and socio-environmental activities that encourage societal and community interaction by working with diverse sectors and professions with the ultimate goals of contributing to social change and promoting health .In 2002, as part of its social mission in collaboration with the Gonçalo Moniz Institute of the Oswaldo Cruz Foundation (IGM-Fiocruz), EBMSP established the Integrative Multidisciplinary HTLV Center (CHTLV) to provide an array of health services for PLHTLV and their families. CHTLV was inspired by the Interdisciplinary HTLV Research Group (GIPH) of Belo Horizonte (state of Minas Gerais), which initiated activities in January 1997 . The cenConsidering that the general guidelines providing for the clinical follow-up of HTLV patients have been well established by the Department of STD, AIDS and Viral Hepatitis , here weThe Singular Therapeutic Project is a set of proposals detailing articulated therapeutic approaches for individuals, families or groups, arising from collective discussions among members of an interdisciplinary team . In accoIn accordance with the nursing process, patient sociodemographic data and current and past health history are obtained during the nursing consultation . Nurses Since most individuals who receive care at CHTLV are unaware of the eligibility to receive benefits from the Brazilian Welfare System due to HTLV-related disabilities, social workers play an important role in counseling PLHTLV on the obtainment of such benefits.Since its inauguration in 2002, a total of 2,169 HTLV-infected patients have been seen at CHTLV, approximately less than 50% of whom are evaluated regularly. Of these, 2,145 (98.9%) are cases of HTLV-1 infection, while 24 (1.1%) individuals were diagnosed with HTLV-2. Almost all patients (98%) reside in the city of Salvador , are aged between 5 and 93 years [mean: 49.8 (SD 15.9)], and 70.3%  are female. Most (84.6%) self-reported black or brown skin color, 73% had less than 8 years of schooling, and half earned the equivalent of one Brazilian monthly wage (∼US$200). Thus, most are considered low-income and possess low education levels. The PLHTLV seen at CHTLV face significant difficulties to attend consultations due to physical limitations and/or lack of access to public transportation.The clinical profile observed in CHTLV patients is similar to that of another center in Salvador , 56, 57:Based on the high frequency of KCS and the difficulty in diagnosing dry eye, an algorithm, using low-cost and minimally invasive tools, for the diagnosis of this disease was developed. Briefly, a sequence of tests in three stages was proposed rather than performing all tests simultaneously. First, patients are submitted to the Ocular Surface Disease Index (OSDI) questionnaire and the tear breakup time test (TBUT). If results from both tests are normal, a diagnosis of KCS can be excluded. However, if the OSDI and/or TBUT results are abnormal, a Schirmer I test is performed, with positivity confirming KCS diagnosis. Lastly, in patients with an indeterminate diagnosis , Rose Bengal staining must then be performed . UnexpecWith respect to HTLV-1 transmission, a pilot project at CHTLV identified a prevalence of 53.5% (146/273) for family aggregation of HTLV-1 infection. Greater TSP/HAM prevalence was observed among index cases. Probable sexual and vertical transmission was estimated at 44.4 and 22.3%, respectively. Index and family cases reported a history of 1–5 partners, with no condom use prior to the diagnosis of HTLV-1. However, enhanced adherence to the use of condoms was described after HTLV-1 diagnosis . A studyHuman T-cell lymphotropic virus type-1-infected patients seen at CHTLV were estimated to face an overall 2.6 times greater relative risk of developing tuberculosis . The preStudies investigating depression in our cohort identified a prevalence ranging from 34.1 to 38.0% , 43. WhiThe rate of HTLV-1 infection in pregnant women in the state of Bahia ranks among the highest in Brazil, with several evaluations reporting prevalence between 0.84 and 1.05% , 74–76 bCHTLV currently serves 178 pregnant/postpartum women. The sociodemographic characteristics of the pregnant women are similar to those of the other women seen at CHTLV, with the exception of younger age [mean (SD): 32.7 (±6.5) years]. A preliminary retrospective study conducted between 2003 and 2012 to evaluate the impact of breastfeeding duration on the vertical transmission of HTLV-1 included a representative sample of postpartum women and their 50 children . Of the via dried blood spot testing has been demonstrated as a secure, viable and low-cost method of increasing pregnant women’s access to serological testing during prenatal care, helping to prevent vertical transmission . Screening for HTLV smission . Bahia ssmission .In April 2013, the SESAB recommended the suspension of breastfeeding by HTLV-positive pregnant women and implemented the provision of milk formula for the first year of the newborn’s life. Campaigns in Brazil commonly recommend breastfeeding as a measure to prevent malnutrition, and also portray breastfeeding as an act of maternal love. In addition to stigmatization due to social and cultural pressures, the inability to breastfeed can engender fears about maternal bonding with newborns. On the other hand, continuing to breastfeed may provoke feelings of guilt associated with the responsibility of viral transmission to children. These sensitive issues present challenges to the prevention of mother-to-child transmission. Accordingly, integrative care for HTLV-1-infected women during pregnancy and the postpartum period is crucial to infection prevention across multiple generations. Thus, great effort has been made to deconstruct longstanding beliefs to give new meaning to bottle feeding as a benevolent act of maternal love in order to preserve newborn health. Importantly, no consensus has been reached regarding the benefits of cesarean delivery in preventing the vertical transmission of HTLV. However, immediate clamping of the umbilical cord following delivery has been strongly recommended . In addi1 CHTLV has strived to vaccinate as many PLHTLV as possible in accordance with the priorities established by the Brazilian Ministry of Health . Of the 258 patients who are members of the HTLVida association, 81 were successfully vaccinated with the help of this organization and CHTLV, according to the following criteria: 37 (46%) due to older age, 27 (33%) because of impaired mobility, 8 (10%) due to comorbidities, 6 (7%) were immunosuppressed, and 3 (4%) belonged to specific groups. Of these 81 PLHTLV, we contacted 52 individuals: 41/52 (79%) have HAM/TSP and 10/52 (19%) contracted COVID-19. Vaccinations included: 14 (27%) CoronaVac, 24 (46%) AstraZeneca/Oxford, 11 (21%) Pfizer, and 2 (4%) Janssen. As of February 2022, 24 (46%) HTLV-1 carriers had received two doses, while 25 (48%) had received a booster shot. Side effects were reported by 38 (73%) patients, and the most prevalent symptoms were as follows: 26 (50%) arm pain, 11 (21%) headache, 10 (19%) fever, 9 (17%) excessive tiredness, 6 (11%) chills, and 5 (10%) muscle pain.People living with human T-cell lymphotropic virus have not been considered as a priority group in the COVID-19 vaccination campaign in Brazil. Working together with the advocacy group HTLVida,In accordance with recommendations issued by the Brazilian government due to the COVID-19 pandemic, EBMSP implemented a contingency plan that severely limited in-person patient care at CHTLV. However, the center implemented telehealth consultations to follow patients regularly seen at CHTLV. The remote care provided by EBMSP/CHTLV allowed for the early diagnosis of complications and stimulated adherence to treatment in PLHTLV. In addition, telehealth consultations helped alleviate some psychological distress caused by social distancing . MoreoveThe mostly multidisciplinary research carried out at CHTLV has been designed to alleviate patient suffering and improve QoL . The resTo promote professional development, some actions have been carried out by EBMSP in conjunction with CHTLV, such as: (1) the inclusion of HTLV in research lines of postgraduate courses; to date, 42 and 19 students have obtained their master’s and Ph.D. degrees respectively; (2) the provision of university-based continuing education training for graduate students in psychology, physiotherapy, nursery, and medicine; (3) weekly meetings to discuss HTLV-related cases with undergraduate and graduate students, and professionals working at CHTLV; (4) financial support for professionals and students to participate in scientific meetings, and scholarships for undergraduate students to develop scientific projects.It is known that PLHTLV suffer high rates of discrimination, mainly due to their vulnerable status as reflected by sociodemographic profile, which becomes intensified as a result of HTLV infection. The interdisciplinary group at CHTLV carries out actions that directly or indirectly seek to minimize prejudice and enable better QoL.Psychoeducational therapeutic intervention is offered to patients and their loved ones, providing information and support to better understand and cope with not only their illness, but also social prejudice. Unfortunately, some patients attempt to hide HTLV infection from their relatives, straining family relationships.Our psychotherapeutic group seeks to strengthen PLHTLV in their fight against prejudice, who are often socially isolated from their family, circle of friends and work colleagues. In psychotherapy, they share common experiences, discuss coping strategies and establish, both among themselves and with professionals, bonds reinforced by a qualified support network.In addition, actions are taken to inform the general population about the virus, demystifying false ideas and encouraging preventative measures.Another action carried out by the EBMSP nursing extension program, in partnership with the HTLVida association, promotes artistic activities and art exhibitions as a way to value autonomy and help PLHTLV cope (9a61047e-67fb-4fb5-b426-cd0bba5aa9c5.mp4).via a regional, national and international network.Collaboration with other research centers has allowed us to not only train health care workers and laboratory staff and promote international exchange with students, researchers, and professionals, but also develop relevant scientific research Members of the CHTLV multidisciplinary team have participated in several public engagement sessions with policymakers organized by the city of Salvador and the Bahia state legislative assembly to encourage the adoption of HTLV-related public health measures. Some team members who advocate for PLHTLV also participate on Bahia State Health Secretariat advisory committees.CHTLV organizes biannual meetings with PLHTLV to discuss their needs and report on research findings, and co-sponsors commemorative events with HTLVIDA, a patient support group created in 2010 with almost 600 members. Events include Brazilian HTLV Day (23 March), the Annual Day for the Prevention and Control of HTLV in Salvador (28 September), and World HTLV Day (10 November). In addition, CHTLV/HTLVIDA also promote social events commemorating festivities, such as Christmas and St. John’s Day, which are popular holidays in northeastern Brazil.People living with human T-cell lymphotropic virus require multidisciplinary care involving multiple medical specialties and laboratory and imaging tests of moderate complexity. For instance, due to structural problems associated with the national unified health care system (SUS) in Brazil, the time required to perform exams, such as MRI or CT, which are essential for patient treatment, is prolonged. Moreover, the scarcity of professionals working in some specialized areas requires CHTLV to meet patients’ needs by collaborating with trained physicians who desire to further their academic studies. In the absence of these types of collaborations, patient services risk being disrupted.As PLHTLV suffer from a chronic condition for which there is no specific treatment or cure, they require ongoing care—yet CHTLV is not able to provide care to patients in need . High demand for services may also compromise the continuity of patient care. Improved integration among the various public health services in the state of Bahia will be necessary to ensure the provision of high-quality care to this neglected population.Brazil is a continental country with great ethnic and economic differences that lead to exacerbated health inequities. Taking into account that integrative health care involves not only individuals, but also their families and communities, including biological, psychological, and social health needs, we recognize the enormous challenge that Brazil faces in implementing efficient and humane health care for PLHTLV.We believe that the actions implemented by CHTLV have contributed to (a) improving the quality of healthcare services provided to HTLV carriers; (b) reducing prejudice and discrimination against HTLV carriers; (c) controlling HTLV infection in vulnerable populations throughout the state of Bahia; (d) the generation and dissemination of scientific and technological knowledge about HTLV infection; (e) the implementation and development of public policies aimed at raising HTLV awareness; (f) the training/education of undergraduate/graduate students; and (g) informing public policymakers about the prevalence of infection, in line with the guiding principles of SUS.We strongly recommend that comprehensive efforts to provide healthcare services for PLHTLV include laboratory diagnosis, counseling , clinical and oral health monitoring (for both asymptomatic and symptomatic patients), as well as pharmacological and non-pharmacological therapeutic measures to alleviate suffering and improve QoL in this extremely neglected population.It is important to strengthen the roles played by patient advocacy groups in interacting with patient care providers to better understand patient concerns, as well as to influence the establishment of pertinent public health policies for PLHTLV.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by the Comitê de Ética em Pesquisa em Seres Humanos da Bahiana. Written informed consent from the patients/participants or their legal guardian/next of kin was not required to participate in this study in accordance with the national legislation and the institutional requirements.BG-C designed and supervised the work. BG-C and MFRG wrote the first 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.


Cancer survivors who received gonadotoxic treatment are at‐risk for future infertility and may desire a fertility status assessment (FSA), defined as semen analysis for males and consultation with a reproductive specialist for females. The purpose of this study was to describe the proportion of, and factors associated with, interest in FSA among young adult survivors of childhood cancer.This retrospective single‐institution review included patients with prior gonadotoxic treatment, aged 18–25 years and >1 year from cancer treatment completion, who received a fertility‐focused discussion during survivorship. Documentation of interest in and completion of FSA, worry about infertility, sociodemographic, and clinical characteristics were abstracted from medical records. Multivariable logistic regression was performed to calculate odds ratios (OR) and 95% confidence intervals (95%CI) for factors associated with interest in FSA.N = 259) were on average 19.2 ± 1.2 years at their fertility discussion; 55.6% were male and 57.9% non‐Hispanic white. Interest in FSA was reported by 50.7% of males and 46.1% of females. Factors related to interest in FSA for males and females respectively, included worry about infertility  and ≥2 fertility discussions . Among males, fertility preservation consult/procedure at diagnosis  and high‐risk for infertility  were also associated with interest in FSA.Survivors (Cancer survivors are interested in FSA, particularly those who have had repeated fertility‐focused discussions during survivorship care and who report worry about infertility. For survivors of childhood cancer who are at risk for infertility, a fertility status assessment can provide clarity regarding their current fertility status and options for future biological parenthood. Cancer survivors are interested in fertility status assessment, particularly those who have had repeated fertility‐focused discussions during survivorship care and those who report worry about infertility. Despite interest in a fertility status assessment, few survivors have completed an assessment. It is unclear what proportion of survivors are interested in an FSA during emerging adulthood, while under the care of a pediatric cancer survivorship program. The purpose of this study was to describe the proportion of YASCC interested in and completing, FSA. We also describe clinical and sociodemographic factors associated with FSA interest .2This study utilized a single institution review of clinical data abstracted from electronic medical records among YASCC who received care within the childhood cancer survivor clinic. The multidisciplinary cancer survivor clinic includes oncology, endocrinology, psychology, nursing, and social work providers who see patients beginning around 2 years after completion of their cancer treatment. Survivors are followed until the age of 21 years at which time they transition to adult healthcare; patients over the age of 21 years who have never attended the survivor clinic are eligible for a one‐time visit. Eligibility criteria for this study included (1) prior cancer diagnosis; (2) ≥1 visit to the cancer survivor clinic between 1/1/2017 and 9/30/2019; (3) ≥1 year from completion of cancer treatment; (4) 18.00–25.99 years of age at the time of visit; (5) and a history of receiving gonadotoxic therapy . Survivors were excluded if they had not received a fertility‐focused discussion during their survivorship clinical visit. Additionally, if survivors had received their initial fertility‐focused discussion <18 years of age, they were excluded due to differences in the developmental stage and decision‐making capacity for pursuing FSA. Survivors of brain tumors are followed in a separate survivorship clinic at the study institution and are not represented in this study. Ethical approval for this study was obtained from the Children's Healthcare of Atlanta institutional review board (IRB# 00000416).Consistent with national guidelines, survivors who received gonadotoxic treatment were counseled regarding their risk for infertility with a focused discussion at age 18 years or older. Survivor team endocrinology providers led a discussion of the survivor's risk for treatment‐related infertility, an interpretation of current reproductive hormone levels, and an assessment of fertility‐related worry. The discussion also included options for FSA and family building and interested survivors were provided with a handout with contact information for reproductive clinics where they could complete an FSA; for survivors who expressed interest in an FSA, this was documented in the medical record. Receipt of FSA was documented in the clinical record based on survivor reports at subsequent clinical visits, or if the FSA report was sent to our center, and in some cases, the fertility team conducted follow‐up phone calls with survivors.2.12.1.1Sociodemographic variables were abstracted from participant medical records, including sex, race/ethnicity, insurance type, religion, and zip code. Using zip codes, participants were classified as living in rural or non‐rural areas in accordance with the Health Resources and Services Administration Federal Office of Rural Health Policy.2.1.2n = 126]) versus no expressed interest (documentation that survivor was not interested [n = 46] or absence of documentation regarding interest [n = 87]). Worry about infertility was similarly characterized as expressed worry  versus no expressed worry (documentation that the survivor was not worried [n = 84] or absence of documentation regarding worry [n = 10]). The number of fertility‐focused discussions (1 vs. ≥2), and receipt of FSA  were also abstracted. Fertility consult (n = 9) and fertility preservation procedure (n = 21) prior to gonadotoxic treatment were combined as a single variable (yes vs. no), due to low numbers.The fertility‐focused discussions were documented in the medical record using a standardized note. Interest in FSA was categorized as expressed interest , no fertility discussion during survivorship visits due to patient declining the conversation (n = 6), other reasons (n = 12), or the fertility discussion was deferred due to patient's cognitive status . The final sample consisted of 144 (55.6%) male and 115 (44.4%) female participants.Among 308 participants who met eligibility criteria, 259 84.1%) received a fertility‐focused discussion and were included in this analysis. Reasons for exclusion from the analysis included receiving initial fertility‐focused discussions and/or FSA when the survivor was a minor  years of age and 8.0 (±5.2) years from cancer treatment completion  and females . Univariable analysis of factors associated with interest in an FSA among males and females are presented in Tables S1 and S2, respectively. In a multivariable logistic regression model, factors associated with interest in FSA among males included ≥2 fertility discussions during survivorship , fertility preservation consult and/or procedure before gonadotoxic treatment , worry about future infertility , and high level (compared with low level) of risk for infertility  and worry about future infertility  were related to interest in FSA  participants expressed interest in FSA with similar rates between males n = 7, 50.7% aOnly 4 (2.8%) males and 10 (8.7%) females survivors had completed FSA while being followed in the cancer survivor clinic. Models for FSA completion could not be tested/built due to the relatively low rates of completion.4For survivors of childhood cancer who are at risk for infertility, FSA can provide clarity regarding their current fertility status and options for future biological parenthood. In this study of YASCC, who had received a fertility‐focused discussion during the study window, about half were interested in FSA , although few had completed assessments  before transitioning from our institution to adult healthcare. Prior research has focused on a wide age range of YASCC while our study adds to the literature by demonstrating a large proportion of survivors are interested in FSA during emerging adulthood. Additionally, we utilized an approach that increases the ecological validity of the study findings, as data were abstracted from standardized notes in the medical record and reflected clinical encounters that occurred in the survivorship clinic. Among both males and females, worry about infertility and more than one fertility‐focused discussion during survivorship were associated with interest in FSA; for males, additional factors associated with interest in FSA included being at high‐risk for infertility and having completed a fertility consultation and/or fertility preservation procedure prior to their gonadotoxic treatment.Prior studies of YASCC consistently demonstrate that survivors are interested in information regarding their risk for infertility and education about reproductive health.FSA procedures and results differ for males and females. Males who are at least several years from cancer treatment completion and complete a semen analysis can generally receive a confirmation of fertility or infertility which is unlikely to change as they age. The costs for this procedure are generally <USD $300. For females, results of FSA can provide insight into their current fertility status and reproductive window, but findings are less definitive and females' fertility status changes with age. There is also the potential for higher costs associated with hormonal laboratory evaluation, transvaginal ultrasound to measure antral follicle count, and consultation with the reproductive endocrinologist if insurance coverage is not available. There may be potential adverse psychological consequences for survivors who pursue FSA and receive results that suggest infertility or likelihood of reduced fertility. For example, survivors may experience a sense of loss, regret for not taking part in fertility preservation at the time of cancer diagnosis, or worry regarding perceived social consequences of infertility.For both males and females, two or more fertility‐related discussions were associated with interest in FSA. For some survivors, the information provided in the initial discussion may be overwhelming to process, meaning the survivor may not become interested in FSA until a second or third fertility discussion. For other survivors who are interested in FSA, repeated fertility discussions during survivor clinic visits may be relevant. In our survivor clinic, fertility discussions are generally initially offered at age 18 years, offered again at the time of transition to adult healthcare (age 21 years), and offered/available at any other time if the patient has questions. Cancer survivors have previously expressed strong desires for fertility‐focused education and discussions after cancer treatment,Among males, being at high‐risk for treatment‐related infertility and the receipt of a fertility preservation consult and/or fertility preservation procedure at the time of cancer diagnosis were associated with interest in FSA. Those at high risk for infertility may be aware of this risk and more motivated to determine if their cancer treatment did indeed result in infertility. Treatment‐related infertility risk was not associated with interest in FSA among females. Prior research demonstrates that females can have perceptions of infertility risk that are discordant from their treatment exposures and/or counseling.4.1While this study presents important findings related to interest in FSA among YASCC, there are limitations that need to be considered. The data from this study were obtained from a single site, through chart review of healthcare provider documentation of fertility discussions. It is possible that some survivors received an FSA that was not documented in our medical records. Similarly, survivors' worry about future infertility and interest in FSA may differ if assessed via self‐report, compared with the provider's documentation of their discussion. As these data stem from clinical discussions, we recognize that there may be heterogeneity in how rigorously these outcomes were assessed and documented, with potential variations by the practitioner, which may have led to reporting bias. Worry was dichotomized for this analysis, which does not reflect the range of worry among YASCC. Most survivors in our sample had hormonal laboratory evaluations at some point after cancer treatment and it is possible that those results affected the FSA completion rates. The sample included an underrepresentation of survivors of leukemia and an overrepresentation of survivors of lymphoma, which may have impacted the findings regarding interest in FSA. Survivors of brain tumors were excluded from this study. We excluded survivors who did not receive gonadotoxic treatment, however, prior research has demonstrated that survivors may worry about their fertility status irrespective of their treatment exposures.Future research in this area is warranted to explore the predictive validity of interest in FSA in relation to FSA completion. As reproductive health has become a focus of cancer survivorship care, there is a need to develop interventions to assist survivors in accessing reproductive health services to optimize options for parenthood. Within the context of emerging adulthood, survivors are experiencing changes and transitions in healthcare, school, work, and family life. This is a time period with the potential for high reproductive health needs, and developmentally appropriate interventions are needed to support survivors' decisions to pursue FSA.In conclusion, this study highlights an important discrepancy in reproductive healthcare among YASCC. Most survivors are worried about future infertility and half report interest in FSA, yet few have received an FSA. Future research is needed to explore barriers to accessing reproductive health services and decision‐making related to FSA. Counseling survivors regarding their risk for infertility and providing referrals for FSA may benefit survivors in emerging adulthood.Brooke Cherven: conceptualization, data curation, formal analysis, funding acquisition, investigation, methodology, project administration, resources, software, supervision, validation, visualization, writing ‐ original draft, and writing ‐ review and editing.Rebecca Williamson Lewis: conceptualization, data curation, formal analysis,, methodology, project administration, software, validation, visualization, writing ‐ original draft, and writing ‐ review and editing.Megan Pruett: conceptualization, data curation, writing ‐ original draft, and writing ‐ review and editing.Lillian Meacham: conceptualization, data curation, funding acquisition, investigation, methodology, validation, visualization, writing ‐ original draft, and writing ‐ review and editing.James Klosky: conceptualization, data curation, formal analysis, funding acquisition, investigation, methodology, resources, validation, visualization, writing ‐ original draft, and writing ‐ review and editing.This Investigator‐Initiated Nursing Research Project was supported by the Children's Oncology Group National Clinical Trials Network Operations Grant . Research reported in this publication was also supported by the National Institute Of Nursing Research of the National Institutes of Health under Award Number K23NR020037. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.The authors report no conflicts of interest.This study received ethical approval from the Children's Healthcare of Atlanta institutional review board (IRB# 00000416), and was conducted in accordance with the Declaration of Helsinki.Table S1‐S2Click here for additional data file.


With the progress of medical technology and renovated conception of fertility, the prospective studies and practice of fertility preservation are drawing more and more attention from medical workers. With the largest population of over 1.4 billion, China makes the experience accumulated in fertility preservation efforts even more relevant. This article summarizes China’s experience and shares it with the world to promote the healthy development of fertility preservation.This study was based on multiple Chinese expert consensuses on fertility preservation issued in 2021 and the current national regulations and principles, compared with the latest advice and guidelines issued by global reproductive authorities such as the ASRM and ESHRE. Summarize the experience and reflection of Chinese scholars in the process of fertility preservation.This study reports on the current situation of fertility preservation in China, sharing the Chinese experience gained in the process of development, and offering Chinese reflections on worrying issues.Fertility preservation is a medical and social issue of reproductive health security, which is conducive to the sound development of the world population and social production. As fertility rates are declining globally, ART has been designed to address the fertility needs of couples with infertility since the very beginning, and as medical technology advances and ideology on childbearing changes, fertility protection and preservation based on prospective fertility appeal are becoming increasingly important to ART practitioners and reproductive biologists. Since the first fertility preservation guidelines were published by American Society of Clinical Oncology in 2006 , fertiliStarting from the current status of fertility preservation in China and with reference to the latest recommendations and guidelines published by global professional authorities such as ASRM, SART, ESHRE, and ISFP, this article discusses China’s decisions and reflections on the development of fertility preservation, summarizes China’s experience, and shares it with the world to promote the healthy development of fertility preservation.Population issues are increasingly attracting the attention of the international community, and the birth rate in China was 8.52‰ in 2020, falling below 10‰ for the first time, while the natural population growth rate was only 1.45‰ in the same period . The ave6/ml to 72.55 × 106/ml [6/ml to 21 × 106/ml [In addition, the incidence of infertility/sterility worldwide shows an increasing trend, which has become a global medical and social problem. There is a lack of national large-scale data on the prevalence rate of infertility/sterility in China. In regional studies, the prevalence rate of infertility among women of reproductive age varies, but still shows an overall increasing trend. According to recent reports, the prevalence rate of infertility among women of childbearing age in Henan and Liaoning provinces of China reached 24.6% and 13.4%, respectively, in 2021 , 11. The× 106/ml ; the humIn summary, China’s population growth rate and fertility rate are decreasing year by year, the trend of delaying the childbearing age is serious, and the prevalence rate of infertility/sterility is increasing year by year, which has a long-lasting and far-reaching impact on social production and economic development. It is imperative to implement measures to protect and preserve the fertility of Chinese men and women of childbearing age as soon as possible and to promote long-term balanced population development.At present, China lacks special statistics on fertility preservation, so this section will indirectly reflect the current situation of fertility preservation in China through the existing data. As of December 2020, there were 536 medical institutions and 27 human sperm banks approved to carry out ART in Chinese mainland. Guangdong has 56 ART medical institutions, ranking first among mainland provinces. Then there are 33 in Jiangsu, and 32 in Shandong and Henan, respectively. However, there is only 1 ART medical institution in Tibet, followed by 2 in Qinghai and Ningxia, respectively  and there are 24 institutions with more than 4000 cycles (accounting for 5.9%) in 2020. 341 institutions with cycles ranging from 100 to 4,000 are the majority (accounting for 84.3%), but there are 36 institutions with less than 100 cycles (accounting for 8.9%)  freezing, reproductive tissue freezing, and sub-tissue freezing Fig. , with vaOvarian/testicular tissue cryopreservation of patients, one of the most prominent clients in fertility preservation efforts, will inevitably carry residual cancer cells, posing a risk of malignancy recurrence for future in situ transplantation. Although rare, there is still a greater risk of recurrence for certain tumors with a high risk of recurrence . For cerCryobiology is the core foundation of fertility preservation technology, and we have experienced the progress from slow freezing to cryopreservation. Currently, vitrification is used as the major method of cryopreservation. Although milestones have been achieved in its development, it is still inconclusive whether it will pose long-term effects over time. The use of frozen embryo transfer, this cohort study includes slow freezing and cryopreservation, was significantly associated with an increased risk of cancer in children, mainly including leukemia and neurological tumors . In 2020In China, there is still no national plan for the establishment of “egg banks,” and only some qualified fertility centers have set up “egg freezing and storage banks” on an exploratory basis. Throughout the world, the first human egg bank was established in Argentina in 2002, and in 2013, the ASRM issued guidelines that no longer considered oocyte freezing a research project, but as a routine ART technique . Since tCompared with women, the medical technology, the high volume of male spermatozoa per discharge, the non-invasive sperm acquisition, the mature technology of sperm freezing and resuscitation, the social background, the high acceptance of the public, the low ethical controversy, and the low space for commercialization, have laid a good foundation for the pioneering work of male fertility protection and the establishment of “sperm bank.” China’s first human sperm bank was established in 1981, and after more than 40 years of development, China’s human sperm banks have formed a relatively standardized and perfect management system to ensure the smooth development of male fertility preservation in China.The most influential journal in the field of reproductive medicine in China published a special feature in 2022, highlighting the medical and social importance of establishing “egg banks” in China , but at PAR is a special phenomenon in the development of ART, including both planned PAR and unplanned PAR—planned PAR means that the deceased had fertility preservation during his or her lifetime, i.e. there was already frozen sperm, eggs, or embryos; unplanned PAR is a sudden accident or death due to an acute illness occurs to the deceased, and his or her spouse or parents request gametes from his or her remains for ART . The latThe first PAR case was publicly reported in China in 2001, which occurred in the case of Zheng Xueli in Zhejiang Province, where the court denied the client’s request for postmortem sperm retrieval for PAR from the husband of the condemned prisoner . The firIn Chinese judicial practice, there is an increasing emphasis on the “Best Interests of the Child” as an important theoretical support for adjudication. For PAR offspring, it is inevitable that they will have the single-parent or reconstituted family as a special growing environment with the psychological and mental aspects of the possible irreversible harm, which is unfair to the offspring and irresponsible of the court decision. At the same time, how to identify the legal status of PAR offspring and protect their rights and interests needs to be urgently resolved. Moreover, PAR inevitably violates the dignity of the deceased and infringes on the deceased’s right to informed consent; and the remains and tissues of the deceased do not belong to general objects and are not legal objects, so they cannot be disposed of by their relatives and friends as the subject of inheritance.In China, due to the deep-rooted traditional concept of “carry on the family line,” especially after the accidental death of a male in a family, his parents and spouse will have an extremely strong claim to the PAR. In addition to considering the interests of the PAR applicants, the rights and interests of the deceased should be fully respected, and the interests of the PAR descendants should be fully evaluated. Countries with similar cultural backgrounds and social environments should be equally cautious when conducting PAR. This not only poses new challenges to medical technology, laws, and regulations, but also places higher demands on civilization.A recommendation published by the ASRM is worth noting. A patient has strongly requested that her physician transfer embryos into her body when conception is almost impossible. The ASRM believes that there are valid and reasonable reasons to support the patient’s decision to transfer at his or her own discretion in order to safeguard his or her reproductive right, and defines this type of embryo transfer as “compassionate transfer” . SimilarStem cells have the ability to self-renew, proliferate, and differentiate into a variety of functional cells with directional differentiation. As the researches of stem cells continue to advance, fertility preservation is gradually explored and practiced, and various stem cells have been found to induce spermatogenesis, including SSCs, MSCs, ESCs, and iPSCs Fig. .Fig. 5VaSSCs can be isolated from testicular tissue for cryopreservation and can be differentiated into spermatozoa by autologous transplantation or in vitro culture when the time is right –54. MSCsWith the support of national policies and increasing awareness of ART, and with the growing social demand for fertility preservation, fertility preservation in China has much room for development. Harboring the aim of promoting China’s progress together with the rest of the world and contributing China’s strength and wisdom to human fertility preservation, we actively learn about key issues in fertility preservation, discuss hot topics that have attracted widespread social attention, share China’s experiences and solutions, and present China’s concerns and thoughts.


Chemotherapy regimens and radiotherapy are common strategies to fight cancer. In women, these therapies may cause side effects such as premature ovarian insufficiency (POI) and infertility. Clinical strategies to protect the ovarian reserve from the lethal effect of cancer therapies needs better understanding of the mechanisms underlying iatrogenic loss of follicle reserve. Recent reports demonstrate a critical role for p53 and CHK2 in the oocyte response to different DNA stressors, which are commonly used to treat cancer. Here we review the molecular mechanisms underlying the DNA damage stress response (DDR) and discuss crosstalk between DDR and signaling pathways implicated in primordial follicle activation. The mammalian ovary is the primary female reproductive organ involved in oocyte maturation and in the synthesis and secretion of sex steroid hormones, estrogens and progesterone, which are crucial for female fertility . The corThe activation of mammalian primordial follicles consists in an irreversible mechanism where a primordial follicle leaves the quiescence status and starts oocyte growth . FolliclRecent studies have demonstrated the adverse effects of cancer therapies on the reproductive potential of young women . ExposurChemotherapy drugs can damage the ovary through different mechanisms . In micePI3K-PTEN-AKT-FOXO3-mTOR and the Hippo signaling pathways are considered two primary regulators of primordial follicle activation at the time of the recruitment into the growing follicle pool (both pathways are represented in The activation of the PI3K complex is triggered by the binding of receptor tyrosine kinase (KITL) ligand, released by pre-granulosa cells (GCs), to its receptor (KIT) on the oocyte. Once activated, PI3K modifies phosphatidylinositol-4,5-bisphosphate (PIP2) into phosphatidylinositol-3,4,5-triphosphate (PIP3). The latter drives the recruitment of both pyruvate dehydrogenase kinase 1 PDK1 and Akt at the membrane, where PDK1 phosphorylation activates Akt. In turn, Akt moves into the nucleus where it phosphorylates the forkhead box O3 (FOXO3) transcription factor . In primAkt is also able to phosphorylate the tuberous sclerosis 2 protein (TSC2), which forms a heterodimer complex with tuberous sclerosis 1 protein (TSC1). In cells, the TSC1/TSC2 complex negatively regulates a target of rapamycin complex 1 (mTORC1) , the latAs mentioned before, the Hippo signaling pathway -Warts (Wts)-Hippo (Hpo) (SWH) pathway) has a crucial role in controlling cell growth and organ size . The HipAs mentioned above, the ovarian reserve is composed of a finite pool of primordial follicles whose number is established before birth. This ovarian reserve represents the capability to generate offspring . In womeCancer treatments with higher genotoxic potential, such as alkylating drugs, cyclophosphamide (Cy), and platinum-based complexes create DNA adducts that interfere with cell cycle progression and replication ,45. ChemStudies in mouse models indicate that TAp63alfa is expressed in primordial oocytes . Excess Ovarian follicle development and oocyte competence are coordinated by a very complex interplay between signaling pathways of different types of cells  forming the ovary. Granulosa cells play a crucial role in follicle growth, they produce hormones, and, through points of contact between cell membranes, they exchange signaling molecules and nutrition with the oocyte. Nutrition, oxygen, and hormone supply are also ensured by the vascular system, which implies the involvement of signaling pathways regulating the angiogenesis process in follicular activation. In mice, the transcription factor FOXO3 is essential in follicular development, but other key signaling molecules underlying follicle activation still remain elusive. Recent reports described multiple signaling pathways underlying primordial follicle activation and maintenance of follicle quiescence ,70. Chem


Alternative iodine remediation technologies include microbial reduction of IO3– to iodide (I–) and microbial methylation of I– to volatile gases. The metal reduction pathway is required for anaerobic IO3– respiration by the gammaproteobacterium Shewanella oneidensis. However, the terminal IO3– reductase and additional enzymes involved in the S. oneidensis IO3– electron transport chain have not yet been identified. In this study, gene deletion mutants deficient in four extracellular electron conduits  and DMSO reductase (ΔdmsB) of S. oneidensis were constructed and examined for anaerobic IO3– reduction activity with either 20 mM lactate or formate as an electron donor. IO3– reduction rate experiments were conducted under anaerobic conditions in defined minimal medium amended with 250 μM IO3– as anaerobic electron acceptor. Only the ΔmtrA mutant displayed a severe deficiency in IO3– reduction activity with lactate as the electron donor, which suggested that the EEC-associated decaheme cytochrome was required for lactate-dependent IO3– reduction. The ΔmtrA-ΔdmsEF triple mutant displayed a severe deficiency in IO3– reduction activity with formate as the electron donor, whereas ΔmtrA-ΔmtrDEF and ΔmtrA-ΔSO4360 retained moderate IO3– reduction activity, which suggested that the EEC-associated dimethylsulfoxide (DMSO) reductase membrane-spanning protein DmsE, but not MtrA, was required for formate-dependent IO3– reduction. Furthermore, gene deletion mutant ΔdmsB  and wild-type cells grown with tungsten replacing molybdenum  in defined growth medium were unable to reduce IO3– with either lactate or formate as the electron donor, which indicated that the DmsAB complex functions as an extracellular IO3– terminal reductase for both electron donors. Results of this study provide complementary genetic and phenotypic evidence that the extracellular DMSO reductase complex DmsAB of S. oneidensis displays broad substrate specificity and reduces IO3– as an alternate terminal electron acceptor.Microbial iodate (IO To tranmembrane , (ii) exompounds , and (iiompounds .3–-reducing microorganisms indicated that nitrate (NO3–) reductase may catalyze the reduction of IO3– as an alternative electron acceptor , and IdrA represented a novel clade within the dimethylsulfoxide (DMSO) reductase family -dependent IrdA for dissimilatory IO3– reduction    and decaand MtrC . MtrC isand MtrC . Other pase CcmB  and the eidensis . On the -factors .3– reduction by S. oneidensis with lactate, but not formate, as the electron donor  protein secretion and metal reduction genes encoding the outer membrane MtrAB module of the EEC MtrCAB are required for IOon donor . However to IO3– .S. oneidensis genome harbors three additional gene clusters that encode the EECs MtrDEF, DmsEFAB, and SO4357-4360 (mtrB mutant under Fe(III) citrate-reducing conditions (S. oneidensis reduces IO3– with separate lactate (MtrAB)- and formate-dependent EEC paralogs that deliver electrons extracellularly to IO3– .In addition to MtrCAB, the 357-4360 . MtrCAB 357-4360 . Further357-4360 . For exa357-4360 , while M357-4360 . MtrDEF 357-4360 . While tnditions . These f3– reductase of S. oneidensis. The experimental strategy consisted of the following steps: (i) construction of three EEC paralog mutants via the deletion of mtrDEF, dmsEF, and SO4360 in a ΔmtrA mutant host strain, and subsequent testing for IO3– reduction activity; (ii) replacement of Mo with tungsten (W) in defined growth medium and tests for IO3– reduction activity; and (iii) construction of DMSO reductase mutant ΔdmsB (deficient in DMSO reductase protein DmsB) and tests of ΔdmsB for IO3– reduction activity with formate and lactate as the electron donor.The main objective of this study was to identify the IOShewanella oneidensis strains were routinely cultured aerobically at 30°C in lysogeny broth (LB) . IO3– reduction rate experiments were conducted under anaerobic conditions in M1 minimal medium  or chloramphenicol (25 μg ml–1) was amended to the appropriate growth medium for the selection of deletion mutant or the maintenance of recombinant plasmid vector pBBR1MCS  [primers D1/D2 and D3/D4  and mobilized into S. oneidensis ΔmtrA . Single crossover integrations were verified using PCR with primers flanking the recombination region (TF/TR) and were resolved from the genomes by plating on LB agar lacking NaCl and containing sucrose [10% (wt/vol)]. The in-frame deletion strains  were verified by PCR with primers TF/TR  of 0.02 was incubated at 30°C for 24 h. Subcultures were centrifuged at 4,000 rpm for 30 min, resuspended in 10 ml of M1 growth medium amended with 10 mM formate and incubated aerobically at room temperature for 8 h. The preconditioned cells were inoculated in the 30-ml serum bottles at an initial OD600 of 0.1 in M1 growth medium amended with either 40 mM DMSO or 250 μM IO3– and 10 mM formate and incubated anaerobically via continuous sparging with 100% high-purity (hydrated) N2 gas. Cultures were incubated at room temperature with gentle stirring under anaerobic conditions maintained by continuous sparging with high-purity hydrated N2 gas. At preselected time points, OD600 was measured and IO3– concentrations were determined using the IO3–-triiodide formation method . Absorbance at 352 nm was measured with a UV spectrophotometer  within the first 3 min of reaction time. IO3– concentrations were determined from a previously generated calibration curve.The extent of IOe method . CultureS. oneidensis employs periplasmic and outer membrane proteins other than MtrAB to deliver electrons to IO3–, three additional EEC gene mutant strains  were constructed in a ΔmtrA mutant host strain. A ΔmtrA gene deletion background was selected since ΔmtrA retained wild-type IO3– reduction activity with formate as electron donor  were determined with either lactate or formate as the electron donor.To test the hypothesis that on donor . Mtr proon donor . To avoi3– reduction activities when compared to wild-type rates  IOon donor .mtrAΔmtrDEF and ΔmtrAΔSO4360 mutant strains provided with formate as the electron donor displayed moderately impaired (between 51 and 70% of wild-type activity) IO3– reduction activities when compared to wild-type rates  with the wild-type strain , corresponding to only 17 and 11% of the rates measured after growth in Mo-containing defined minimal medium   . These fS. oneidensis employs DmsAB as the IO3– terminal reductase, we generated a ΔdmsB gene deletion mutant. In a previous study, mutant strain ΔdmsB was unable to grow anaerobically with DMSO as a terminal election acceptor  grew at wild-type rates with DMSO as electron acceptor , outer membrane β-barrel protein (OMP), and extracellular terminal reductase (ETR) (S. oneidensis EECs are involved in the reduction of Fe(III), Mn(IV), Mn(III), flavins, and DMSO , Mn(IV), Mn(III), and flavin reduction systems. The overexpression of MtrD restores wild-type Fe(III)-citrate reduction rates to an ΔmtrA mutant  . S. oneiand DMSO . Severaland DMSO . S. oneid SO4360 . MtrA isA mutant . DmsE isA mutant , but parA mutant . SO4360 function .S. oneidensis ETRs are involved in Fe(III) or DMSO reduction and include the decaheme c-type cytochromes MtrC, MtrF, and OmcA. MtrF and OmcA functionally replace MtrC  (3– reduction by S. oneidensis with lactate (but not formate) as the electron donor ] as the terminal electron acceptor  . SO4359, SO4359) . The Mtron donor . Similaracceptor . Based oS. oneidensis IO3– reduction system was proposed -dependent proposed . In thisependent . In the ependent , which fein DmsF . DmsA anein DmsF . The extof DmsAB , is unliS. oneidensis IO3– reduction system utilizes MtrAB and does not require DmsEF, but does require DmsAB as the IO3– terminal reductase. DmsAB is evolutionarily unrelated to MtrC, MtrF, or OmcA. Although the Mtr respiratory pathway is modular, MtrAB has not been reported to transfer electrons to extracellular reductases apart from MtrC, MtrF, or OmcA. However, previous work which tested all possible combinations of Mtr paralogs  system of Pseudomonas sp. strain SCT and Denitromonas sp. IR-12 homologs that display amino acid sequence homology to respiratory arsenite oxidase; however, both systems require molybdopterin coordinating Mo (3– reduction, yield details important to the biogeochemical cycling of iodine in marine systems, and provide information crucial to the development of alternative bioremediation technologies for the treatment of radioactive iodine-contaminated subsurface environments.The extracellular IOating Mo . ResultsThe original contributions presented in the study are included in the article/H-DS, YT, JM, and RM performed the experiments. H-DS, YT, and TD wrote the manuscript. All authors had given approval to the final version of the manuscript.Any opinion, findings, and conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of the US Department of Energy Office of Environmental Management.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.


Calcium influx can be stimulated by various intra- and extracellular signals to set coordinated gene expression programs into motion. As such, the precise regulation of intracellular calcium represents a nexus between environmental cues and intrinsic genetic programs. Mounting genetic evidence points to a role for the deregulation of intracellular calcium signaling in neuropsychiatric disorders of developmental origin. These findings have prompted renewed enthusiasm for understanding the roles of calcium during normal and dysfunctional prenatal development. In this Review, we describe the fundamental mechanisms through which calcium is spatiotemporally regulated and directs early neurodevelopmental events. We also discuss unanswered questions about intracellular calcium regulation during the emergence of neurodevelopmental disease, and provide evidence that disruption of cell-specific calcium homeostasis and/or redeployment of developmental calcium signaling mechanisms may contribute to adult neurological disorders. We propose that understanding the normal developmental events that build the nervous system will rely on gaining insights into cell type-specific calcium signaling mechanisms. Such an understanding will enable therapeutic strategies targeting calcium-dependent mechanisms to mitigate disease. Summary: In this Review, we describe calcium-dependent mechanisms regulating early neurodevelopmental events in the cortex and provide evidence for calcium deregulation as a convergence point in the etiology of neurodevelopmental disorders. Calcium, which lies at the hub of multiple signal transduction pathways, is uniquely situated to transduce dynamic biological inputs into distinct cell behaviors  are a primary route of calcium influx in developing neural cells  in respoAnother major source of calcium influx is store operated calcium entry (SOCE), which is activated upon endoplasmic reticulum (ER) calcium store depletion in response to extracellular signals  reviewe. SOCE isCalcium signaling involves both the sensing and transduction of extracellular calcium , as wellCalcium influx through different calcium-permeable channels engages distinct signaling cascades , in partLocal translation of calcium signaling components can also influence calcium dynamics in specific cellular compartments, such as neuronal dendrites . MoreoveXenopus nervous system identified two types of spontaneous calcium transients contributing to distinct aspects of spinal neuron differentiation  are two major sources of calcium influx in developing neural cells  reviewe. Growingv1.2 . In the v1.2 . This biDifferentiation in the embryonic cortex is accompanied by global splicing changes . MoreoveCACNA1C (encoding the pore-forming subunit of the VGCC Cav1.2), for example, is extensively spliced to generate channels with distinct properties , which are mutated in the syndromic autism spectrum disorder (ASD) Timothy Syndrome (TS), is developmentally controlled  Gene database for studies implicating regulators of calcium signaling in the etiology of neurodevelopmental disorders. We restrict our analysis , are implicated in ASD   and chromosomal abnormalities in loci containing γ-aminobutyric acid receptor (GABAR) genes (e.g. 15q11-13) have been associated with neurodevelopmental phenotypes , proliferative neural stem cells that populate the cortex with neurons and astrocytes  proliferation  and intermediate progenitor cells (IPCs) , which gSpontaneous calcium rises are dependent on internal calcium stores, as they persist in the absence of extracellular calcium but are eliminated upon ER calcium depletion . VGCC acGrowth factors are important cell cycle regulators , which yields elevated chloride concentrations in embryonic neural cells (+-Cl− co-transporter KCC2 (SLC12A5) induces a developmental switch in GABA activity from depolarizing to hyperpolarizing  and kainate (KA) receptors are expressed in VZ/SVZ cells . Four machannels . AlthougThe neurotransmitters serotonin (5-HT) and acetylcholine (Ach) also induce NSPC calcium transients. Maternally-derived 5-HT is reported to regulate proliferation in the developing brain ; howeverIt is possible that agonist-induced SOCE and depolarization-induced calcium entry represent mechanisms that antagonistically regulate NSPC proliferation. Understanding the coordination of different modes of calcium entry and how their interplay impacts proliferative NSPCs is thus essential. Moreover, as calcium influx via VGCCs and SOCE can contribute to calcium oscillations, it will be crucial to determine how cellular responses are encoded in patterns of calcium transients and how these dynamics enable signaling specificity in NSPCs.Proliferative RGCs undergo interkinetic nuclear migration (INM) – dynamic somatic movements in phase with the cell cycle , undertaking saltatory tangential migration to reach the cortex , canonically bound by brain derived neurotrophic factor (BDNF), has been linked to embryonic cortical neuroblast migration  Fig. 3)Fig. 3). igration .Trkb, the number of calbindin-positive interneurons that reach the cortex is attenuated  and Gad67 (Gad1) do not display major cortical malformations (Gad67 knockout mice (ARs (e.g. taurine) have thus been proposed to control neuronal migration  activation by endogenous ligands  is also depolarizing in cortical neuroblasts, eliciting calcium rises that may modulate migration . AlthougGRIN2B, encoding GluN2B, is highly expressed in postmitotic embryonic neurons, whereas GRIN2A, encoding GluN2A, is expressed in embryonic RGCs and neurons after birth -expressing interneurons . PostnatAdcyap1) and transduced via calcium has been identified as a complementary mechanism involved in the onset of astrogliogenesis (Gfap) gene (Dream (Kcnip3) fails to induce astrocyte differentiation, but this can be rescued by JAK/STAT activation, suggesting that calcium-regulated DREAM signaling works in parallel to JAK/STAT signaling during astrogliogenesis  gene . These dap) gene . PACAP sogenesis .v1.2 loss of function results in impaired OPC proliferation, axon-OPC interactions and myelination (Embryonic oligodendrocyte precursor cells (OPCs) initially populate the cortex through two waves of migration from the ganglionic eminences . A thirdlination , 2016. Llination . What inlination , such OPin vitro  and microglia . DissectAs we have reviewed here, the coordinated spatiotemporal regulation of calcium signaling directs cellular behaviors that underlie early cortical development, including aspects of NSPC function. Calcium is a key mediator of activity-dependent gene expression, serving as a hub linking environmental cues to the cytoskeleton, metabolic pathways and other biochemical cascades. Underscoring these crucial developmental roles, genetic studies reveal that mutations in genes encoding calcium signaling modulators contribute to the pathophysiology of neurodevelopmental disorders. There remain, however, open questions about how activity and calcium-dependent processes are regulated at a cell type-specific level and how this regulation coordinates cellular behaviors during corticogenesis. In other cell and tissue types, properties of intracellular calcium dynamics have been linked to activation of specific downstream transcription factors. It is thus tempting to hypothesize that distinct cell type-specific patterns of electrical signals and calcium elevations in the embryonic cortex may subserve specific developmental roles. A natural corollary to this hypothesis is that disrupting calcium signaling and patterned electrical activity in specific cortical populations during development may promote neurodevelopmental disease.Much remains to be elucidated about how calcium signaling is initiated and transduced in developing cortical cells. Titration of calcium signaling is not only achieved through dynamic regulation of ion channels and signaling proteins but also via coordination of extrinsic cues, including maternal hormones and metabolic regulation , which iIn primary glioma specimens from adult human patients, stem-like glioma cells display molecular signatures reflecting neural stem and progenitor cell (NSPC) identity , and theCalcium signaling deregulation has been implicated in neurodegenerative disorders, including Alzheimer's disease (AD)  and Down syndrome (DS) have increased risk of developing AD and related dementias early in life . AlthougDetermining how disease-relevant mutations affect intracellular signaling and cell type-specific developmental behaviors across space and time (reviewed by 10.1242/develop.198853_sup1Supplementary informationClick here for additional data file.


Drosophila models of neurological disease contribute tremendously to research progress due to the high conservation of human disease genes, the powerful and sophisticated genetic toolkit, and the rapid generation time. Fragile X syndrome (FXS) is the most prevalent heritable cause of intellectual disability and autism spectrum disorders, and the Drosophila FXS disease model has been critical for the genetic screening discovery of new intercellular secretion mechanisms. Here, we focus on the roles of three major signaling pathways: BMP, Wnt, and insulin-like peptides. We present Drosophila FXS model defects compared to mouse models in stem cells/embryos, the glutamatergic neuromuscular junction (NMJ) synapse model, and the developing adult brain. All three of these secreted signaling pathways are strikingly altered in FXS disease models, giving new mechanistic insights into impaired cellular outcomes and neurological phenotypes. Drosophila provides a powerful genetic screening platform to expand understanding of these secretory mechanisms and to test cellular roles in both peripheral and central nervous systems. The studies demonstrate the importance of exploring broad genetic interactions and unexpected regulatory mechanisms. We discuss a number of research avenues to pursue BMP, Wnt, and insulin signaling in future FXS investigations and the development of potential therapeutics. Drosophila genome contains ∼ 70% conserved homologs of human disease genes, which have been repeatedly proven to mediate equivalent functions in similar cells and tissues  to reduce transcripts at specific times and in defined cells. For neurological disease models, the developing Drosophila central nervous system (CNS) has been extensively characterized at the level of individually-identified neural stem cells and neurons (Drosophila glutamatergic neuromuscular junction (NMJ) offers superior imaging and electrophysiological access that has proven invaluable in modeling numerous disease states  as the founding Wnt ligand (Drosophila insulin-like peptide (ILP) secretion and signaling  gene  is the leading neurodevelopmental disorder causing inherited intellectual disability , often aR1) gene , leadingR1) gene . A few rR1) gene . ClinicaR1) gene . In mammR1) gene . Only FM defects , includi defects , defecti defects , and imp defects . Recent FMR1, FXR1, and FXR2) mutants. For example, FXR2 deficiency mice exhibit inhibition of BMP signaling through upregulation of the secreted BMP-binding Noggin, which normally functions in preventing BMP ligands from binding to their receptors, resulting in aberrant neural progenitor cell (NPC) proliferation and differentiation within the hippocampal dentate gyrus (DG)  . Acting  DG-NPCs . BMP sig DG-NPCs . Gene extrans-synaptic BMP signaling regulates synaptic structure and function at the Drosophila larval glutamatergic NMJ (futsch (human MAP1B) mRNA translation by repressing mRNA-bound FMRP function, thus up-regulating synaptic growth (BMPR2 mRNA (Drosophila Wit homolog) to inhibit full-length isoform translation, thus causing accumulation of the noncanonical BMP pathway component Lin11/Isl1/Mec3 domain kinase 1 (LIMK1) within neurons  mRNA to stabilize the transcripts in muscle (coracle (cora) mRNA to restrict translation of this glutamate type II A receptor (GluRIIA) anchoring actin scaffold  + DrosophilaMad) signaling serve to link mitochondrial and Wnt signaling regulatory networks (see below). Mechanistically, augmented phospho-SMAD1/5/9 (pSMAD1/5/9) binds to the tumor suppresser p53 protein, thus preventing p53 degradation from forming complexes with ubiquitin ligase murine double minute 2 (MDM2) (Bax-mediated apoptotic pathway in BMP type 1A receptor (BMPR1a) mutant mice  . Consequant mice . To inhiant mice . While Fant mice , we hypoDrosophila FXS stem cells, FMRP also suppresses the insulin-like receptor (InR) via LIN-28, an RNA-binding protein required for the translation of insulin-like growth factors  activation  signaling in FXS originated from elevated phosphatase and tensin (PTEN), target of rapamycin (TOR), phosphoinositide 3-kinase (PI3K), and activated protein kinase B (Akt) in FXS model and patient neurons , consist factors . Drosophtivation . Geneticn adults . Moreoven adults , suggestDrosophila FXS model, FMRP regulation of ILP signaling is involved in CNS development. In Drosophila progenitor stem cells (neuroblasts) and subsequently, in developing glia, FMRP sequentially limits the reactivation of larval brain neuroblasts by inhibiting ILP signaling  and subsequent glial engulfment and clearance of axons  glial phagocytosis mechanism of neural clearance  levels are also significantly reduced on glial membranes, correlating with the delayed developmental clearance of the PDF-tri neurons. Furthermore, constitutively activating glial InRs in dfmr1 mutants restores normal neuron clearance, showing that FMRP-dependent glial InR activation is required for phagocytosis  and developmental-stage  dependent  clock neurons, FMRP is required to mediate removal from the early adult la brain . In the learance . In dfmrocytosis . Moreoveocytosis . Note thependent . It is nependent , which aependent . FMRP isependent . It willDrosophila Wingless (Wg), whose developmental role was identified in the famous segment polarity screen  and evenness interrupted (Evi)  . Wnt reced (Evi) . Wnt siged (Evi) . In FXS ed (Evi) . FMRP bied (Evi) . This stIn the mouse FXS model, pharmacological inhibition of GSK3β significantly improves hippocampus-dependent learning by rescuing neurogenesis and neuronal maturation defects , furtherDrosophila FXS model, FMRP regulates trans-synaptic Wnt signaling to modulate glutamatergic NMJ larval synaptogenesis  glypican Dally-like protein (Dlp) acting as a Wg co-receptor, as well as the transmembrane HSPG syndecan (Sdc), are both highly elevated at the NMJ synaptic terminal , a proteinase that cleaves extracellular proteins, to modulate larval synaptic structure and function by modulating secreted Wnt signaling (dfmr1 mutants exhibit upregulation of MMP1 and MMP1 proteolytic enzymatic activity surrounding presynaptic boutons at the glutamatergic NMJ. Drosophila only has two MMPs (secreted MMP1 and the GPI-anchored MMP2), with the secreted protease specifically affected by FMRP. In the mouse adult FXS model, secreted MMP7/9 are likewise positively upregulated out of at least 23 MMPs, correlated with Wnt signaling differences  and drive synaptic bouton formation  in different developmental stages, particularly in the nervous system. New discoveries suggest that FMRP occupies core roles linking BMP, ILP, and Wnt regulatory networks that mediate neurogenesis, gliogenesis, synaptogenesis, and glial functions during neural circuit remodeling. To study synaptogenesis and neurotransmission function regulated by this FMRP-dependent signaling, the ormation . This noormation . While tormation , we do ntic FMRP , the mectic FMRP , it woulDrosophila motor neurons, insulin signaling also negatively regulates presynaptic neurotransmitter release via the FOXO-dependent regulation of the eukaryotic initiation factor 4e binding protein (4eBP) translational inhibitor  scaffold recruitment to the muscle subsynaptic reticulum (SSR)  . With thum (SSR) , it is pelevated , but we Drosophila BMP ligands  have been well characterized in the CNS, but likely show a similarly robust distribution. In both mouse and Drosophila, multiple reports demonstrate that BMP signaling plays essential roles in communication between neurons and glia in development, in neural remodeling after injury, and during aging (Drosophila FXS model, FMRP is required in neurons, but not glia, for glial phagocytosis during brain clock circuit remodeling (Drosophila BMPR2 homolog Wit mRNA to inhibit translation (Multiple FMRP-dependent secreted signals likely mediate intercellular communication between neurons and glia in brain development and circuit remodeling . In the ng aging . Howevermodeling . FMRP binslation , suggestnslation . In thisnslation , and redDrosophila InRs in ensheathing glia and astrocyte-like glia are required for neuronal clearance following injury (Drosophila, signaling downstream of activated glial InRs promotes Akt phosphorylation, which is essential for Draper phagocytosis receptor expression (The role of FMRP-dependent insulin and Wnt signaling in neuron-glia communication has just begun to be studied. Neurally secreted ILPs are suspected to activate glial engulfment for neuronal clearance . Howeverg injury  as well g injury . In miceg injury . In Drospression . These dpression . It willpression . Moreovepression . Perhapspression . Similarpression . BMP-Wntpression . Neverth


Giardia duodenalis is a common parasitic diarrheal agent in humans, especially in developing countries. The aim of this study was to investigate the prevalence and multilocus genetic characterization of G. duodenalis in patients with diarrhea and animals in northeastern China, and to assess the epidemiological role of animals in the transmission of human giardiasis.G. duodenalis by PCR and sequencing of the SSU rRNA gene. All G. duodenalis-positive specimens were subtyped by PCR and sequencing of the bg, tpi, and gdh genes. To detect additional mixed infections of different assemblages, assemblage A/B/E-specific PCRs were performed to amplify the tpi gene.A total of 1739 fecal specimens from 413 diarrheal patients and 1326 animals comprising 16 mammal species were collected in Heilongjiang Province of China and screened for SSU rRNA gene determined the prevalence of G. duodenalis  in diarrheal patients, with a peak in minors aged 5–17 years, and identified assemblages A and B. MLG-AII and MLG-B1 were obtained based on concatenated nucleotide sequences of the bg, tpi, and gdh genes, with MLG-AII being identical to a cat-derived isolate reported previously. By sequence analysis of the SSU rRNA gene, G. duodenalis was detected in 214 (16.14%) animals belonging to 11 mammal species, with the prevalence ranging from 1.69 to 53.85%, and assemblages A to G were identified. Sequence analysis of the bg, tpi, and gdh genes from 46 specimens produced 31 MLGs, including MLG-AI (n = 1), MLG-B2–B8 (n = 18), and MLG-E1–E23 (n = 27).Sequence analysis of the G. duodenalis in diarrheal patients enhances consciousness of detecting G. duodenalis in clinical practice and emphasizes the importance of health education in local inhabitants, especially in the age group of 5–17 years. The identification of seven assemblages (A to G) and 33 MLGs reveals genetic heterogeneity of G. duodenalis in the investigated areas. Due to insufficient homology data on the zoonotic transmission of G. duodenalis, the precise epidemiological role that animals play in the transmission of human giardiasis needs to be assessed by more large-scale molecular epidemiological investigations of local humans and animals.The finding of The online version contains supplementary material available at 10.1186/s13071-022-05269-9. Giardia duodenalis . In fact, giardiasis has been included in WHO’s Neglected Diseases Initiative since 2004 ), and housekeeping genes (triosephosphate isomerase [tpi] and glutamate dehydrogenase [gdh]) ) . Based oubtyping . To date A and B .G. duodenalis infection in humans are common, with approximately 28.5 million cases of G. duodenalis infection estimated in 2005 [G. duodenalis infection have been carried out in five provinces/municipalities, with three assemblages  being identified [G. duodenalis in diarrheal patients in China is unclear. In northeastern China’s Heilongjiang Province, G. duodenalis infection has ever been documented in humans in about 70% of counties/cities based on the first national epidemiological investigation conducted in 1988–1992 [G. duodenalis infection in humans. To investigate the prevalence and multilocus genetic characterization of G. duodenalis in patients with diarrhea and in animals, and to assess the epidemiological role of animals in the transmission of human giardiasis, a molecular epidemiological study of G. duodenalis was carried out in patients with diarrhea and various animals by multilocus genotyping of G. duodenalis isolates.In China, epidemiological investigations and case reports on  in 2005 . Since gentified , 27, 28.988–1992 . Since tn = 150), Central Hospital of Yichun Forestry Administration Bureau in Yichun (n = 222), Jianhua Hospital in Qiqihar (n = 26), Jidong Central Hospital in Jixi (n = 8), and Kangying Hospital in Suihua (n = 7) , Suihua (n = 366), Shuangyashan (n = 22), Qiqihar (n = 31), Daqing (n = 18) and the Greater Khingan Mountains (n = 207)  were collected from humans and animals (one each) from October 2010 to May 2021 in Heilongjiang Province, China. Of these, 413 human fecal specimens were from patients with diarrhea at least three times each day and who were outpatients in five hospitals: the Second Affiliated Hospital of Harbin Medical University in Harbin . The procedures and reagents used in the present study were provided by the manufacturer. DNA was finally eluted in 200 μl of AE elution buffer and stored at −20 °C until polymerase chain reaction (PCR) analysis.Genomic DNA of G. duodenalis by nested PCR amplification of an approximately 290 base pairs (bp) of the SSU rRNA gene, and were identified to the assemblage level as previously designed by Appelbee et al. [G. duodenalis-positive specimens at the SSU rRNA locus were further analyzed and identified to the sub-assemblage level by nested PCR amplifications of the bg, tpi, and gdh genes. The fragments of the bg (~ 510 bp), tpi (~ 530 bp), and gdh (~ 530 bp) genes were amplified as previously designed by Lalle et al., Sulaiman et al., and Caccio et al., respectively [tpi gene as previously designed by Geurden et al. and Levecke et al. [G. duodenalis assemblage A isolate was used as positive controls in PCR tests to amplify the SSU rRNA, bg, tpi , and gdh genes. DNA of assemblages B and E from a human and a sheep was used as positive controls in assemblage B/E-specific PCR tests to amplify the tpi gene, respectively. DNase-free water was used as a negative control in each PCR test. All secondary PCR products were separated by 1.5% agarose gel electrophoresis, following staining with GelStrain , observed, photographed, and recorded on a Gel Doc™ EZ Gel Documentation System .All DNA specimens were screened for the presence of e et al. . All G. ectively –33. In ae et al. , 35. DNAhttp://www.ncbi.nlm.nih.gov/blast/) and the program Clustal X 1.83  to determine the assemblages and sub-assemblages of G. duodenalis.All positive secondary PCR products of the expected size were sequenced by Comate Bioscience Company Limited  using the respective secondary PCR primers on an ABI PRISM 3730xl DNA Analyzer using the BigDye Terminator v3.1 Cycle Sequencing Kit . The nucleotide sequence accuracy was confirmed by two-directional sequencing and by sequencing two more new PCR products if necessary for some DNA specimens in which novel nucleotide sequences were acquired. Nucleotide sequences obtained in the present study were aligned and analyzed with each other and reference sequences of each locus downloaded from GenBank using the Basic Local Alignment Search Tool (BLAST)  were included in the multilocus genotyping analysis. Nucleotide sequences of each isolate at the three analyzed loci were concatenated (bg-tpi-gdh) to form one MLG.All the specimens successfully amplified and sequenced at the three loci , OM001842 and OM001843 (tpi), and OM001857 to OM001859 (gdh); animal-derived G. duodenalis isolates—OM001822 and OM001825 to OM001839 (bg), OM001840, OM001841 and OM001844 to OM001855 (tpi), OM001856 and OM001860 to OM001879 (gdh).The representative nucleotide sequences obtained in the present study were deposited in the GenBank database under the following accession numbers: human-derived χ2) tests implemented in SPSS  19.0. were used to compare differences in prevalence among different age groups. Odds ratios (ORs) with 95% confidence intervals (CIs) were calculated for the occurrence of G. duodenalis in humans. Differences were considered statistically significant at a P-value of < 0.05.Fisher’s exact test and Pearson Chi-square  of human fecal specimens and identified assemblages A (n = 5) and B (n = 19). Giardia duodenalis was distributed in three age groups: 9.78% (22/225) in the minors aged 5–17 years, 1.20% (1/83) in the middle-aged adults aged 36–60 years, and 2.08% (1/48) in the older adults aged > 60 years. By χ2 tests, a significant difference in prevalence was observed only between the age groups of 5–17 and 36–60 years  , and three specimens were successfully amplified and subtyped, respectively. All DNA specimens did not have successful PCR amplifications at the tpi locus using general primers. By sequence analysis, sub-assemblages AII (n = 2) and B1 (n = 1) were identified at the bg locus, while sub-assemblages AII (n = 1) and B1 (n = 1) at the tpi locus, and sub-assemblages AII (n = 1), B1 (n = 1) and B2 (n = 1) at the gdh locus. One gdh nucleotide sequence (B1) was not described previously but had 100% similarity with the nucleotide sequences from a human (MG736274), a rabbit (KP635094), and a monkey (MK952603) at the amino acid level , 336 bp (tpi for A) or 384 bp (tpi for B), and 498 bp (gdh), two MLGs (MLG-AII and MLG-B1) were produced  of fecal specimens from 11 mammal species. Sheep (Ovis aries) , cattle (Bos taurus) , and rabbits (Oryctolagus cuniculus)  had much higher prevalence than the other animals—red deer (Cervus elaphus) , sika deer (Cervus nippon) , reindeer (Rangifer tarandus) , raccoon dogs (Nyctereutes procyonoides) , cats (Felis catus) , brown rats (Rattus norvegicus) , dogs (Canis lupus familiaris) , and pigs (Sus scrofa domesticus) . Assemblages A to G were identified in the animals. The host distribution of the different assemblages is shown in Table Sequence analysis of the partial n 16.14% 14/1326 oG. duodenalis at the SSU rRNA locus were subjected to PCR amplifications of the bg, tpi, and gdh genes, and 73, 88, and 74 DNA specimens were successfully amplified and subtyped, respectively. Analysis of 73 bg gene sequences identified sub-assemblage AI (n = 3), B2–B4 (n = 25), D (n = 1), E1–E10 (n = 42), and G (n = 2). The two bg gene sequences of assemblage G were identical to each other. At the tpi locus, 19 G. duodenalis-positive specimens were successfully sequenced using general primers, and all of them belonged to assemblage E. Eighty-eight G. duodenalis-positive specimens were successfully sequenced using assemblage-specific primers, and were identified as sub-assemblages AI-1 and AI-2 (n = 7), B2–B4 (n = 32), and E1–E9 (n = 51), which covered the above 19 positive specimens , B3–B5 (n = 25), D1 and D2 (n = 2), E1–E14 (n = 41), and G (n = 3). The three gdh gene sequences of assemblage G were identical to each other  , 31 MLGs were obtained: MLG-AI (n = 1), MLG-B2 to MLG-B8 (n = 18) and MLG-E1 to MLG-E23 (n = 27)  + E (at the SSU rRNA locus) in one cow, A (at the tpi locus) + E (at the SSU rRNA and bg loci) in one sheep, A (at the tpi locus) + E (at the SSU rRNA locus) in one sika deer, and C (at the SSU rRNA locus) + D (at the gdh locus) in one dog, as well as A + E in one cow and one sheep by assemblage-specific nested PCR assays targeting the tpi gene  and individuals 16–75 years (22.2%) [G. duodenalis infection could be attributed to poor hygiene and play habits of this age group. All 24 diarrheal patients with giardiasis in the present study claimed no experience of direct contact with animals. However, previous studies have considered that close contact with animals was one of the possible risk factors related to G. duodenalis infection. Possible zoonotic transmission has been observed between humans and hoofed animals  in Brazil, Egypt, Canada, Vietnam, and India [Giardia duodenalis has a wide animal host range, and it has been identified in various mammal species. In the present study, G. duodenalis was found in 16.14% of investigated animals, and was identified for the first time in pigs, reindeer, red deer, sika deer, and brown rats in Heilongjiang Province of China, defining the host distribution of G. duodenalis across this region. In the present study, we only identified G. duodenalis with zoonotic potential in brown rats, and not Giardia muris, which is specific mainly to rodents belonging to the family Muridae [In the investigated diarrheal patients, a prevalence peak was observed in the 5–17 age group (9.78%). A similar age pattern of  (22.2%) . In Chin (22.2%) . The abond India , 39, 40. Muridae .SSU rRNA gene, assemblage B  was observed to be more prevalent than assemblage A  in diarrheal patients. As the major assemblages A and B causing human giardiasis, assemblage B was responsible for more infections (4083 cases) than assemblage A (2833 cases), based on molecular analysis of 7211 specimens of G. duodenalis from two review articles [SSU rRNA gene revealed assemblage constituents of G. duodenalis in different animal species: D and E in cattle; A and E in sheep, reindeer, and red deer; E in pigs and sika deer; C in dogs and raccoon dogs; F in cats; B in rabbits; G in brown rats. Because 95.79% (205/214) of animal-derived isolates identified as assemblages A to F are pathogenic for humans, the animals infected with G. duodenalis might pose a risk of zoonotic transmission of human giardiasis, and were of public health importance. The identification of assemblage D in cattle was unexpected. Assemblage D is mainly found in canine animals and occasionally seen in other animals, including cattle. To date, only two studies have reported assemblage D in cattle in China and the UK [G. duodenalis cysts of assemblage D may have been successfully transmitted to cattle through contaminated water or food.By sequence analysis of the articles , 42. In d the UK , 44. Basbg, tpi, and gdh genes exhibited genetic heterogeneity within assemblages A, B, and E in the investigated areas. Two sub-assemblages were identified with AI in animals and AII in humans. To date, identification of the genetic variations in nucleotide sequences within assemblage A at the bg, tpi, and gdh loci has led to the designation of sub-assemblages AI, AII and AIII. They all appear to have different host specificity: AI commonly found in animals, AII largely seen in humans, and AIII mainly detected in wild mammals and rarely in companion animals, cattle, and red deer [G. duodenalis isolates, suggesting that concerns about the zoonotic transmission of G. duodenalis should be directed toward these animals, especially cattle, sheep, reindeer, sika deer, and rabbits infected with zoonotic assemblages A and B. The multilocus genotyping analysis further supported the above presumption of zoonotic transmission of assemblages A and B in the investigated areas. No identical MLGs were found between humans and animals in the present study. Thus, the epidemiological role of animals as reservoir hosts in the transmission of G. duodenalis to humans needs to be assessed in the future. For assemblage E, commonly found in hoofed mammals, no identical MLGs were found in cattle and sheep, reflecting limited cross-species transmission of G. duodenalis occurring between animals living on the same farms or grazing in the same pastures.Sequence analysis of the red deer . By homoG. duodenalis in patients with diarrhea and animals in northeastern China’s Heilongjiang Province. The finding of G. duodenalis in diarrheal patients enhances awareness regarding the detection of G. duodenalis in clinical practice and emphasizes the importance of health education in local inhabitants, especially in the age group of 5–17 years. Seven assemblages (A to G) and 33 MLGs were obtained—A and B (two MLGs) in humans and A to G (31 MLGs) in animals—revealing genetic heterogeneity of G. duodenalis in the investigated areas. Assessment of possible zoonotic transmission of G. duodenalis was based on homology data mostly at the single locus level and rarely at the multilocus level, and no identical MLGs were observed between the investigated humans and animals. The precise epidemiological role that animals play in the transmission of human giardiasis needs to be assessed by further large-scale molecular epidemiological investigations of local humans and animals.The present study describes the prevalence and multilocus genetic characterization of Additional file 1: Table S1. Homology analysis of the bg gene of G. duodenalis isolates at the nucleotide and amino acid levels. Table S2. Homology analysis of the tpi gene of G. duodenalis isolates at the nucleotide and amino acid levels. Table S3. Homology analysis of the gdh gene of G. duodenalis isolates at the nucleotide and amino acid levels.