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UMass Chan Medical School has three graduate schools: the T.H. Chan School of Medicine, the Morningside Graduate School of Biomedical Sciences and the Tan Chingfen Graduate School of Nursing. The collections in these communities archive and share research from UMass Chan students of all three schools, including doctoral dissertations, master's theses, published journal articles, research projects, capstone presentations and conference materials.

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Recently Published

  • Regulation of Lipolysis by 14-3-3 Proteins on Human Adipocyte Lipid Droplets

    Yang, Qin (2024-04-11)
    Lipid droplets (LDs) in adipocytes are pivotal for systemic lipid metabolism, serving as storage centers during nutritional surplus and as sources of fatty acids when energy is needed. These LDs react to hormonal stimuli like catecholamines and insulin, and their impaired response can lead to dysregulated lipolysis, lipotoxicity, and an increased risk of metabolic diseases. The specific mechanisms behind lipid release in human adipocytes remain largely unexplored. This study aims to elucidate the control of lipid mobilization in human adipocytes. We utilized advanced techniques to generate and differentiate primary progenitor cells on a large scale. Employing proximity labeling with enhanced ascorbate peroxidase (APEX2), we identified the interactome of perilipin 1 (PLIN1), a key LD component protein, under various lipolytic states. Through LC-MS/MS, we discovered 70 proteins interacting specifically with PLIN1. This includes PNPLA2 and LIPE, vital for regulated triglyceride hydrolysis, and four 14-3-3 protein family members (YWHAB, YWHAE, YWHAZ, YWHAG), which are known to regulate diverse signaling pathways. Our functional studies revealed that YWHAB is essential for maximal cyclic adenosine monophosphate (cAMP)-stimulated lipolysis, as its CRISPR-Cas9-mediated knockout mitigates lipolysis through a mechanism independent of insulin signaling. In summary, our use of proximity labeling not only comprehensively mapped the LD proteome in human adipocytes but also unveiled new regulatory mechanisms in adipocyte lipolysis control, specifically involving 14-3-3 proteins.
  • A Pilot Clinical Trial of an Informatics-Informed Mobile Transitional Care Program: the Paramedic Assisted Community Evaluation after Discharge (PACED) Intervention

    O'Connor, Laurel (2024-04-03)
    Introduction: Early rehospitalization of frail older adults after hospital discharge is detrimental to patients and hospital systems. Implementing effective strategies to execute a feasible and effective transitional care plan is challenging. Mobile integrated health (MIH) programs, which deploy mobile assets into the community to care for patients, may present a possible solution to facilitating effective transitional care back to home environments after hospitalization. However, there have been few previous studies investigating MIH models for transitional care delivery. The objective of this project is to assess the implementation and effectiveness of an informatics-supported paramedic-led MIH transitional care program for frail older adults. Methods: Patients 65 and older preparing for discharge from the hospital with an eFrailty index of 0.24 or greater were enrolled to participate in a structured post-discharge transitional home visit conducted by community paramedics within 72 hours of discharge. Demographic and clinical information, as well as healthcare utilization patterns, were recorded at enrollment and 30 days after the index hospitalization. Additionally, a separate control group of patients that were screened for the intervention but excluded due to geographical location of residence were aggregated and their electronic health record data including demographical and outcomes data was abstracted. Categorical group comparisons were conducted using chi-square tests and continuous variables group comparisons were conducted using the Kruskal–Wallis equality-of-populations rank test. Crude and adjusted binomial regressions were used for comparative outcomes. Results: In total 100 subjects were enrolled in the intervention (median age 81, 64% female) and 47 were included in the control group (median age 80, 55.2% female). The recruitment rate was 18.0%. The complete intervention protocol was completed and documented by paramedics for 90 (90.0%) patients. The crude and relative risk of 30-day rehospitalization was decreased in the PACED group compared to the control (RR=0.40, CI 0.19-0.84, p=0.03). There was a non-significant trend toward decreased risk of 30-day ED visits (RR=0.61, CI=0.37-1.37, p=0.23). Paramedics identified medication errors in 34 (34.0%) of the participants; the errors were remediated during the visit in 31 (91.2%). Additionally, 67 (67.0%) of subjects screened positive for high fall risk and 7 subjects (7.0%) screened positive for delirium. Conclusions: This pilot study of MIH intervention transition care program programs was feasible with high protocol fidelity and yields preliminary evidence that the intervention results in a decreased risk of rehospitalization in frail older adults.
  • Vocal learning-associated convergent evolution in mammalian proteins and regulatory elements

    Wirthlin, Morgan E; Schmid, Tobias A; Elie, Julie E; Zhang, Xiaomeng; Kowalczyk, Amanda; Redlich, Ruby; Shvareva, Varvara A; Rakuljic, Ashley; Ji, Maria B; Bhat, Ninad S; et al. (2024-03-29)
    Vocal production learning ("vocal learning") is a convergently evolved trait in vertebrates. To identify brain genomic elements associated with mammalian vocal learning, we integrated genomic, anatomical, and neurophysiological data from the Egyptian fruit bat (Rousettus aegyptiacus) with analyses of the genomes of 215 placental mammals. First, we identified a set of proteins evolving more slowly in vocal learners. Then, we discovered a vocal motor cortical region in the Egyptian fruit bat, an emergent vocal learner, and leveraged that knowledge to identify active cis-regulatory elements in the motor cortex of vocal learners. Machine learning methods applied to motor cortex open chromatin revealed 50 enhancers robustly associated with vocal learning whose activity tended to be lower in vocal learners. Our research implicates convergent losses of motor cortex regulatory elements in mammalian vocal learning evolution.
  • Enhancing Safety and Efficacy of Genome Editing In Vivo with Compact Cas9 and Guide Chemical Modification

    Zhang, Han (2024-03-25)
    Clustered regularly interspaced short palindromic repeats and CRISPR-associated proteins (CRISPR/Cas) have revolutionized science and medicine. However, applying the CRISPR/Cas system for in vivo therapeutic genome editing remains challenging due in part to delivery obstacles. Adeno-associated virus (AAV) vectors remain one of the most promising vehicles for gene therapies, particularly in extrahepatic tissues. Nevertheless, employing AAV for in vivo CRISPR/Cas genome editing presents difficulties, including limited packaging capacity, prolonged expression, redosing complexities, modest multiplexing capability, dose-related toxicity, and immunogenicity. This thesis addresses AAV limitations in delivering CRISPR/Cas in vivo in two ways. First, I focus on developing a novel delivery modality in which the CRISPR RNA (crRNA) is delivered in the form of a naked oligonucleotide, separate from AAV-expressed effector and trans-activating crRNA (tracrRNA). I find that a short, fully stabilized oligonucleotide (a ‘protecting oligo’) can significantly enhance the potency, stability, and uptake of a heavily chemically modified crRNA. The establishment of AAV/crRNA co-delivery modality offers a route towards transient editing activity, target multiplexing, guide redosing, and vector inactivation. Second, I use a compact Nme2Cas9 adenine base editor (Nme2-ABE) to engineer a single AAV delivery system for base editing in vivo. The capability to deliver a base editor via a single AAV holds the potential to enhance safety by minimizing immune responses, mitigating dose-related toxicity, and reducing manufacturing complexity.
  • Introducing Telomere-Dysfunction Induced Cellular Senescence in iPSC-Based Modeling Of Neurodegenerative Diseases

    Neherin, Kashfia (2024-03-06)
    Investigation of disease-associated cellular changes in vitro has leaped forward significantly with the innovation of induced pluripotent stem cell (iPSC) technology. However, resetting the epigenetic landscape and aging clock during reprogramming results in iPSC-differentiated cells resembling fetal cell types instead of adult or aged cells. The lack of cellular aging in iPSC-based models presents a significant drawback in the investigation of age-associated diseases such as Alzheimer’s disease. My thesis aims to introduce proper cellular aging to improve iPSC-based modeling of neurodegeneration. Toward this goal, I created an inducible system to trigger senescence in iPSC-based cell models, as recent studies showed senescence playing a crucial role in aging and neurodegeneration. I utilized CRISPR-interference (CRISPRi) to suppress telomere repeat factor 2 (TERF2), a significant component of the telomere-protecting Shelterin complex. I demonstrated that suppression of TERF2 in iPSCs robustly activated DNA damage response (DDR), p53/p21 signaling, and cellular senescence in an inducible and synchronized manner. The inducible approach allows temporal control of senescence activation throughout differentiation from iPSCs to desired cell types. I applied the CRISPRi-TERF2 approach to iPSC-differentiated neural progenitor cells (NPCs) and showed that suppression of TERF2 efficiently activated DDR, p53/p21 signaling, and cellular senescence in differentiated NPCs. This inducible model of cellular senescence generated in this study will enable the investigation of cellular senescence using isogenic comparisons in the progression of age-associated neurodegeneration and improve disease modeling with a proper cellular aging context to facilitate drug discovery.
  • Silencing Parkinson's risk allele Rit2 sex-specifically compromises motor function and dopamine neuron viability

    Kearney, Patrick J; Zhang, Yuanxi; Liang, Marianna; Tan, Yanglan; Kahuno, Elizabeth; Conklin, Tucker L; Fagan, Rita R; Pavchinskiy, Rebecca G; Shaffer, Scott A; Yue, Zhenyu; et al. (2024-02-23)
    Parkinson's disease (PD) is the second most prevalent neurodegenerative disease and arises from dopamine (DA) neuron death selectively in the substantia nigra pars compacta (SNc). Rit2 is a reported PD risk allele, and recent single cell transcriptomic studies identified a major RIT2 cluster in PD DA neurons, potentially linking Rit2 expression loss to a PD patient cohort. However, it is still unknown whether Rit2 loss itself impacts DA neuron function and/or viability. Here we report that conditional Rit2 silencing in mouse DA neurons drove motor dysfunction that occurred earlier in males than females and was rescued at early stages by either inhibiting the DA transporter (DAT) or with L-DOPA treatment. Motor dysfunction was accompanied by decreased DA release, striatal DA content, phenotypic DAergic markers, DA neurons, and DAergic terminals, with increased pSer129-alpha synuclein and pSer935-LRRK2 expression. These results provide clear evidence that Rit2 loss is causal for SNc cell death and motor dysfunction, and reveal key sex-specific differences in the response to Rit2 loss.
  • Racial and Ethnic Disparities in Use of Colorectal Cancer Screening Among Adults With Chronic Medical Conditions: BRFSS 2012-2020

    Castañeda-Avila, Maira A; Tisminetzky, Mayra; Oyinbo, Atinuke G; Lapane, Kate L (2024-02-22)
    Introduction: People with chronic conditions and people with colorectal cancer (CRC) may share common risk factors; thus, CRC screening is important for people with chronic conditions. We examined racial and ethnic differences in the use of CRC screening among people with various numbers of chronic conditions. Methods: We included data on adult respondents aged 50 to 75 years from the Behavioral Risk Factor Surveillance System in 2012 through 2020. We categorized counts of 9 conditions as 0, 1, 2, 3, and ≥4. We classified self-reported CRC screening status as up to date or not. We used Poisson models to estimate adjusted prevalence ratios (APRs) among the different counts of chronic conditions in 4 racial and ethnic groups: Hispanic adults with limited English proficiency (LEP), Hispanic adults without LEP, non-Hispanic Black adults, and non-Hispanic White adults. Results: Overall, 66.5% of respondents were up to date with CRC screening. The prevalence of being up to date increased with the number of chronic conditions. We found disparities among racial and ethnic groups. Hispanic respondents with LEP had lower rates than non-Hispanic White adults of being up to date with CRC screening across all counts of chronic conditions (APR for 0 conditions = 0.67; 95% CI, 0.64-0.71; APR for ≥4 conditions = 0.85; 95% CI, 0.79-0.91). Hispanic respondents without LEP with 0, 1, or 2 conditions were less likely than non-Hispanic White respondents to be up to date with CRC screening. We found no significant differences between non-Hispanic Black and non-Hispanic White respondents. Conclusion: We found disparities among Hispanic BRFSS respondents with LEP, who had lower rates than non-Hispanic White respondents of being up to date with CRC screening, regardless of the number of chronic conditions. Tailored interventions are needed to address these disparities and improve screening rates, particularly among Hispanic people.
  • Persistent False Positive Covid-19 Rapid Antigen Tests

    Herbert, Carly; McManus, David D; Soni, Apurv (2024-02-22)
    Rapid antigen tests for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are effective tools for the diagnosis of acute infection, particularly when used serially. The percentage of rapid antigen tests with false positive results is reported to be less than 1%. However, we have observed persons who repeatedly test positive with rapid antigen tests despite concurrent negative molecular tests; this infrequent phenomenon occurs predominantly among women and persons with autoimmune disorders.
  • Domain-inlaid Nme2Cas9 adenine base editors with improved activity and targeting scope

    Bamidele, Nathan; Zhang, Han; Dong, Xiaolong; Cheng, Haoyang; Gaston, Nicholas; Feinzig, Hailey; Cao, Hanbing; Kelly, Karen; Watts, Jonathan K; Xie, Jun; et al. (2024-02-17)
    Nme2Cas9 has been established as a genome editing platform with compact size, high accuracy, and broad targeting range, including single-AAV-deliverable adenine base editors. Here, we engineer Nme2Cas9 to further increase the activity and targeting scope of compact Nme2Cas9 base editors. We first use domain insertion to position the deaminase domain nearer the displaced DNA strand in the target-bound complex. These domain-inlaid Nme2Cas9 variants exhibit shifted editing windows and increased activity in comparison to the N-terminally fused Nme2-ABE. We next expand the editing scope by swapping the Nme2Cas9 PAM-interacting domain with that of SmuCas9, which we had previously defined as recognizing a single-cytidine PAM. We then use these enhancements to introduce therapeutically relevant edits in a variety of cell types. Finally, we validate domain-inlaid Nme2-ABEs for single-AAV delivery in vivo.
  • Genome-wide kinetic profiling of pre-mRNA 3' end cleavage

    Torres-Ulloa, Leslie; Calvo-Roitberg, Ezequiel; Pai, Athma A (2024-02-16)
    Cleavage and polyadenylation is necessary for the formation of mature mRNA molecules. The rate at which this process occurs can determine the temporal availability of mRNA for subsequent function throughout the cell and is likely tightly regulated. Despite advances in high-throughput approaches for global kinetic profiling of RNA maturation, genome-wide 3' end cleavage rates have never been measured. Here, we describe a novel approach to estimate the rates of cleavage, using metabolic labeling of nascent RNA, high-throughput sequencing, and mathematical modeling. Using in silico simulations of nascent RNA-seq data, we show that our approach can accurately and precisely estimate cleavage half-lives for both constitutive and alternative sites. We find that 3' end cleavage is fast on average, with half-lives under a minute, but highly variable across individual sites. Rapid cleavage is promoted by the presence of canonical sequence elements and an increased density of polyadenylation signals near a cleavage site. Finally, we find that cleavage rates are associated with the localization of RNA polymerase II at the end of a gene, and faster cleavage leads to quicker degradation of downstream readthrough RNA. Our findings shed light on the features important for efficient 3' end cleavage and the regulation of transcription termination.
  • Hydroxyapatite deposition disease, an overlooked differential diagnosis in the emergency department: a case series and review of literature

    Patel, Jay; Tai, Ryan; Sereni, Christopher; Joshi, Ganesh (2024-02-15)
    Hydroxyapatite crystal deposition disease (HADD) poses diagnostic challenges in the emergency department (ED) as it may clinically present similarly to infection and other musculoskeletal conditions. Misdiagnosis often leads to unnecessary treatments and resource over-utilization. This review article provides an overview of HADD in seven patients who presented to the ED secondary to an acute presentation of this disease process. HADD is a prevalent pathology, which commonly involves the shoulder, followed by the hip, elbow, wrist, and knee. Predisposing risk factors, such as diabetes and certain genetic factors, have also been identified. Clinical history and imaging, particularly radiographs, play a vital role in diagnosing HADD, with characteristic calcification patterns observed in different stages of the disease. Conservative nonsurgical therapy is the mainstay of treatment, providing effective symptom relief in over 90% of cases. By recognizing HADD as a crucial differential diagnosis for patients with acute or chronic pain, healthcare resource utilization can be optimized, leading to improved patient care in the ED.
  • Home Diagnostics, Viral Dynamics, and Post-acute Sequelae of SARS-COV-2

    Herbert, Carly (2024-02-09)
    Introduction: The emergence of the novel coronavirus, SARS-CoV-2, has necessitated prompt evaluations of diagnostic technologies, viral dynamics, and long-term complications of COVID-19 to inform clinical and public health strategies. Methods: Using data from the RADx Clinical Studies Core collected from October 2021-February 2022, we evaluated the longitudinal performance of reverse transcriptase polymerase chain reaction (RT-PCR) and antigen-detecting rapid diagnostic tests (Ag-RDT) by day past symptom onset and close-contact exposure and compared performance by sex, age, vaccination status, and variant (Aim 1). We further examined the association between SARS-CoV-2 viral load, BMI, and sex (Aim 2). Lastly, we conducted a follow-up survey in August 2023 regarding Long COVID. We then modeled the relationship between viral clearance of SARS-CoV-2 and Long COVID (Aim 3). Results: RT-PCR and Ag-RDT showed the highest percent positivity two days past symptom onset (RT-PCR: 91.2%; Ag-RDT: 71.1%) and six days past exposure (RT-PCR: 91.8%; Ag-RDT: 86.2%). Performance did not differ by vaccination status, variant, age category, or sex. In males, increasing BMI was associated with higher viral load in a dose-response fashion, and males had significantly lower viral load than females for BMI>29. Lastly, the risk of long COVID with 3-4 symptoms and 5+ symptoms increased by 2.90 times (95% CI: 1.09-7.74) and 4.54 times (95% CI: 1.84-11.2) per viral load slope-unit increase, respectively. Conclusion: Understanding SARS-CoV-2 viral dynamics is critical to identify effective diagnostic strategies for COVID-19, explain differences in COVID-19 outcomes across sex and BMI, and understand mechanistic contributors of Long COVID.
  • Microglia-astrocyte crosstalk regulates synapse remodeling via Wnt signaling [preprint]

    Faust, Travis E; Lee, Yi-Han; O'Connor, Ciara; Boyle, Margaret A; Gunner, Georgia; Badimon, Ana; Ayata, Pinar; Schaefer, Anne; Schafer, Dorothy P (2024-02-09)
    Astrocytes and microglia are emerging key regulators of activity-dependent synapse remodeling that engulf and remove synapses in response to changes in neural activity. Yet, the degree to which these cells communicate to coordinate this process remains an open question. Here, we use whisker removal in postnatal mice to induce activity-dependent synapse removal in the barrel cortex. We show that astrocytes do not engulf synapses in this paradigm. Instead, astrocytes reduce their contact with synapses prior to microglia-mediated synapse engulfment. We further show that reduced astrocyte-contact with synapses is dependent on microglial CX3CL1-CX3CR1 signaling and release of Wnts from microglia following whisker removal. These results demonstrate an activity-dependent mechanism by which microglia instruct astrocyte-synapse interactions, which then provides a permissive environment for microglia to remove synapses. We further show that this mechanism is critical to remodel synapses in a changing sensory environment and this signaling is upregulated in several disease contexts.
  • Bigtools: a high-performance BigWig and BigBed library in Rust [preprint]

    Huey, Jack; Abdennur, Nezar (2024-02-08)
    The BigWig and BigBed file formats were originally designed for the visualization of next-generation sequencing data through a genome browser. Due to their versatility, these formats have long since become ubiquitous for the storage of processed sequencing data and regularly serve as the basis for downstream data analysis. As the number and size of sequencing experiments continues to accelerate, there is an increasing demand to efficiently generate and query BigWig and BigBed files in a scalable and robust manner, and to efficiently integrate these functionalities into data analysis environments and third-party applications. Here, we present Bigtools, a feature-complete, high-performance, and integrable software library for generating and querying both BigWig and BigBed files. Bigtools is written in the Rust programming language and includes a flexible suite of command line tools as well as bindings to Python. Bigtools is cross-platform and released under the MIT license. It is distributed on Crates.io and the Python Package Index, and the source code is available at https://github.com/jackh726/bigtools.
  • Clocks at sea: the genome-editing tide is rising

    Kwiatkowski, Erica R; Rosenthal, Joshua J C; Emery, Patrick (2024-02-08)
    The coastline is a particularly challenging environment for its inhabitants. Not only do they have to cope with the solar day and the passing of seasons, but they must also deal with tides. In addition, many marine species track the phase of the moon, especially to coordinate reproduction. Marine animals show remarkable behavioral and physiological adaptability, using biological clocks to anticipate specific environmental cycles. Presently, we lack a basic understanding of the molecular mechanisms underlying circatidal and circalunar clocks. Recent advances in genome engineering and the development of genetically tractable marine model organisms are transforming how we study these timekeeping mechanisms and opening a novel era in marine chronobiology.
  • Participant Perspectives on the Implementation of a School-Linked Text-Message Intervention to Improve Pediatric Asthma Medication Adherence

    Radu, Sonia; Zarinafsar, Sheerin; Ryan, Grace W; Chainani, Sanjay; Becker, Sarah; Arenas, Juliana; Spano, Michelle A; Shillan, Holly N; Hoque, Shushmita; Sadasivam, Rajani; et al. (2024-02-07)
    Background: Poor adherence to inhaled corticosteroids (ICS) is a significant challenge in pediatric asthma, contributing to health inequities. Text-message reminders for ICS therapy are an evidence-based approach that improves pediatric asthma medication adherence, yet has not been widely adopted into practice, partly due to lack of (1) participant input on design and implementation and (2) use of sustainable community linkages. Remote Asthma Link™ (RAL) seeks to fill this gap as a school-linked text-message intervention wherein parents of children with poorly controlled asthma received daily, 2-way text-message reminders for preventive inhaler use. Responses were shared with school nurses who conducted remote check-ins with families. Enrolled children, largely from underserved backgrounds, experienced improvements in medication adherence and asthma health outcomes. While initial results were promising, we have yet to elicit participant input to refine the protocol for more widespread implementation. Objective: Examine participant perspectives on barriers and facilitators of RAL implementation. Methods: Semistructured interviews were conducted May-June 2022 with intervention participants: 10 parents, 7 school nurses, and 4 pediatric providers (n = 21) until thematic saturation was reached. Interview transcripts were coded using thematic analysis. Results: Several facilitators for RAL implementation were identified, including ease of use and accessibility, personal connection to the school nurse, and receipt of a visual notification for habit formation. Barriers included challenges with school nurses reaching parents, poor understanding of program expectations, and lack of reimbursement structure. Participant-proposed solutions to barriers included utilizing alternate communication methods (eg, social media), educational sessions, and meeting with payors to consider reimbursement models. Conclusion: RAL is a school-linked text-message intervention demonstrating promise in improving outcomes and equity in asthma care. Key implementation facilitators, barriers, and proposed solutions will inform protocol adaptations to promote successful implementation of this and other text-message interventions into clinical practice.
  • Protein and Guide RNA Engineering of a Compact Cas9 for Enhanced Precision Genome Editing

    Bamidele, Nathan (2024-02-06)
    CRISPR-Cas technologies enable robust manipulation of genetic material, and have been instrumental in advancing a wide range of fields across the life sciences. Specifically, with the ability to correct or alter faulty genes, genome editing tools promise to transform the field of genetic medicine. Current CRISPR-based editors [nucleases, base editors (BEs), and prime editors (PE)] can be programed to induce efficient mutagenesis/repair, conversion, and polymerization, respectively. Presently, nucleases - the most clinically advanced genome editors - suffer from inadequate control of genome editing outcomes. Over time, the field has focused on precision editors such as BEs and PEs that do not rely on double-strand breaks and greatly improve the safety and control of genome editing outcomes. Despite these advances, challenges such as targeting scope, accuracy and in vivo delivery represent major hurdles for the therapeutic application of next-generation editing systems such as BE and PE. In this thesis, I focus on alleviating some of the key obstacles associated with effective genome editing by improving the unique properties of a compact Cas9 orthologue (Nme2Cas9 from Neisseria meningitidis). The bulk of my thesis consists of protein engineering efforts to improve the activity and targeting scope of Nme2Cas9-derived editing systems. My later work focuses on the development of chemically stabilized guide RNAs, providing a path to facilitate in vivo delivery in a variety of formats. Overall, the advances presented in this thesis contribute to the versatility of CRISPR-based genome editing systems for a variety of therapeutic and research applications.
  • Microbiota encoded fatty-acid metabolism expands tuft cells to protect tissues homeostasis during infection in the large intestine [preprint]

    Kellogg, Tasia D; Ceglia, Simona; Mortzfeld, Benedikt M; Zeamer, Abigail L; Foley, Sage E; Ward, Doyle V; Bhattarai, Shakti K; McCormick, Beth A; Reboldi, Andrea; Bucci, Vanni (2024-01-31)
    Metabolic byproducts of the intestinal microbiota are crucial in maintaining host immune tone and shaping inter-species ecological dynamics. Among these metabolites, succinate is a driver of tuft cell (TC) differentiation and consequent type 2 immunity-dependent protection against invading parasites in the small intestine. Succinate is also a growth enhancer of the nosocomial pathogen Clostridioides difficile in the large intestine. To date, no research has shown the role of succinate in modulating TC dynamics in the large intestine, or the relevance of this immune pathway to C. difficile pathophysiology. Here we reveal the existence of a three-way circuit between commensal microbes, C. difficile and host epithelial cells which centers around succinate. Through selective microbiota depletion experiments we demonstrate higher levels of type 2 cytokines leading to expansion of TCs in the colon. We then demonstrate the causal role of the microbiome in modulating colonic TC abundance and subsequent type 2 cytokine induction using rational supplementation experiments with fecal transplants and microbial consortia of succinate-producing bacteria. We show that administration of a succinate-deficient Bacteroides thetaiotaomicron knockout (Δfrd) significantly reduces the enhanced type 2 immunity in mono-colonized mice. Finally, we demonstrate that mice prophylactically administered with the consortium of succinate-producing bacteria show reduced C. difficile-induced morbidity and mortality compared to mice administered with heat-killed bacteria or the vehicle. This effect is reduced in a partial tuft cell knockout mouse, Pou2f3+/-, and nullified in the tuft cell knockout mouse, Pou2f3-/-, confirming that the observed protection occurs via the TC pathway. Succinate is an intermediary metabolite of the production of short-chain fatty acids, and its concentration often increases during dysbiosis. The first barrier to enteric pathogens alike is the intestinal epithelial barrier, and host maintenance and strengthening of barrier integrity is vital to homeostasis. Considering our data, we propose that activation of TC by the microbiota-produced succinate in the colon is a mechanism evolved by the host to counterbalance microbiome-derived cues that facilitate invasion by intestinal pathogens.
  • Association of spatial proximity to fixed-site syringe services programs with HCV serostatus and injection equipment sharing practices among people who inject drugs in rural New England, United States

    Romo, Eric; Stopka, Thomas J; Jesdale, Bill M; Wang, Bo; Mazor, Kathleen M; Friedmann, Peter D (2024-01-28)
    Background: Hepatitis C virus (HCV) disproportionately affects rural communities, where health services are geographically dispersed. It remains unknown whether proximity to a syringe services program (SSP) is associated with HCV infection among rural people who inject drugs (PWID). Methods: Data are from a cross-sectional sample of adults who reported injecting drugs in the past 30 days recruited from rural counties in New Hampshire, Vermont, and Massachusetts (2018-2019). We calculated the road network distance between each participant's address and the nearest fixed-site SSP, categorized as ≤ 1 mile, 1-3 miles, 3-10 miles, and > 10 miles. Staff performed HCV antibody tests and a survey assessed past 30-day injection equipment sharing practices: borrowing used syringes, borrowing other used injection equipment, and backloading. Mixed effects modified Poisson regression estimated prevalence ratios (aPR) and 95% confidence intervals (95% CI). Analyses were also stratified by means of transportation. Results: Among 330 PWID, 25% lived ≤ 1 mile of the nearest SSP, 17% lived 1-3 miles of an SSP, 12% lived 3-10 miles of an SSP, and 46% lived > 10 miles from an SSP. In multivariable models, compared to PWID who lived within 1 mile of an SSP, those who lived 3 to 10 miles away had a higher prevalence of HCV seropositivity (aPR: 1.25, 95% CI 1.06-1.46), borrowing other used injection equipment (aPR: 1.23, 95% CI 1.04-1.46), and backloading (aPR: 1.48, 95% CI 1.17-1.88). Similar results were observed for PWID living > 10 miles from an SSP: aPR [HCV]: 1.19, 95% CI 1.01-1.40; aPR [borrowing other used equipment]:1.45, 95% CI 1.29-1.63; and aPR [backloading]: 1.59, 95% CI 1.13-2.24. Associations between living 1 to 3 miles of an SSP and each outcome did not reach statistical significance. When stratified by means of transportation, associations between distance to SSP and each outcome (except borrowing other used injection equipment) were only observed among PWID who traveled by other means (versus traveled by automobile). Conclusions: Among PWID in rural New England, living farther from a fixed-site SSP was associated with a higher prevalence of HCV seropositivity, borrowing other used injection equipment, and backloading, reinforcing the need to increase SSP accessibility in rural areas. Means of transportation may modify this relationship.
  • Investigating the etiologies of non-malarial febrile illness in Senegal using metagenomic sequencing

    Levine, Zoë C; Sene, Aita; Mkandawire, Winnie; Deme, Awa B; Ndiaye, Tolla; Sy, Mouhamad; Gaye, Amy; Diedhiou, Younouss; Mbaye, Amadou M; Ndiaye, Ibrahima M; et al. (2024-01-25)
    The worldwide decline in malaria incidence is revealing the extensive burden of non-malarial febrile illness (NMFI), which remains poorly understood and difficult to diagnose. To characterize NMFI in Senegal, we collected venous blood and clinical metadata in a cross-sectional study of febrile patients and healthy controls in a low malaria burden area. Using 16S and untargeted sequencing, we detected viral, bacterial, or eukaryotic pathogens in 23% (38/163) of NMFI cases. Bacteria were the most common, with relapsing fever Borrelia and spotted fever Rickettsia found in 15.5% and 3.8% of cases, respectively. Four viral pathogens were found in a total of 7 febrile cases (3.5%). Sequencing also detected undiagnosed Plasmodium, including one putative P. ovale infection. We developed a logistic regression model that can distinguish Borrelia from NMFIs with similar presentation based on symptoms and vital signs (F1 score: 0.823). These results highlight the challenge and importance of improved diagnostics, especially for Borrelia, to support diagnosis and surveillance.

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