Discovery of Trypanosoma brucei inhibitors enabled by a unified synthesis of diverse sulfonyl fluorides.

Sets of electrophilic probes are generally prepared using a narrow toolkit of robust reactions, which tends to limit both their structural and functional diversity. A unified synthesis of skeletally-diverse sulfonyl fluorides was developed that relied upon photoredox-catalysed dehydrogenative couplings between hetaryl sulfonyl fluorides and hydrogen donor building blocks. A set of 32 diverse probes was prepared, and then screened against Trypanosoma brucei. Four of the probes were found to have sub-micromolar anti-trypanosomal activity. A chemical proteomic approach, harnessing an alkynylated analogue and broad-spectrum fluorophosphonate tools, provided insights into the observed anti-trypanosomal activity, which likely stems from covalent modification of multiple protein targets. It is envisaged that the unified diversity-oriented approach may enable the discovery of electrophilic probes that have value in the elucidation of biological and biomedical mechanisms.

Assessment of disability and disease burden in neuromyelitis optica spectrum disorders in the CIRCLES Cohort.

Neuromyelitis optica spectrum disorders (NMOSD) comprise autoimmune diseases imposing substantial disability. We compared an NMOSD-targeted disability assessment of mobility, vision, and self-care domains (individually and composite) with the multiple sclerosis-targeted Expanded Disability Status Scale (EDSS) to assess NMOSD disease burden. An overall cohort (n = 505) and a subset of these patients with an enriched dataset (n = 198) were analyzed from the CIRCLES longitudinal, observational database of patients with AQP4-IgG-seropositive or -seronegative NMOSD in North America. Multinomial modeling was used to identify temporal correlates of disability improvement, stability, and worsening. Prior on-study relapse correlated with worsening mobility (OR, 3.08; 95% CI: 1.61-5.90), vision (OR, 3.99; 95% CI: 2.03-7.86), self-care disability (OR, 1.90; 95% CI: 1.07-3.38), and mean composite index disability (OR, 4.20; 95% CI: 1.71-10.34). Higher vision disability was associated with Black race, shorter time on-study, and AQP4-IgG-seropositive status in patients ≥ 18 years (p < 0.05). Disease onset phenotype and sex correlated with pain interference (p < 0.05). These correlates of NMOSD disability were undetected by EDSS. The CIRCLES real-world experience supports the need for NMOSD-specific disability assessment to improve recognition of disease burden, facilitate proactive clinical management, offer insights into resilience, and inform clinical trial design.

Diagnostic Value of Inter-Eye Difference Metrics on OCT for Myelin Oligodendrocyte Glycoprotein Antibody-Associated Optic Neuritis.

BACKGROUND AND OBJECTIVES: The 2022 International Consortium for Optic Neuritis diagnostic criteria for optic neuritis (ON) include optical coherence tomography (OCT). The diagnostic value of intereye difference (IED) metrics is high for ON in patients with multiple sclerosis and aquaporin-4 antibody seropositive neuromyelitis optica spectrum disorders, but unknown in myelin oligodendrocyte glycoprotein antibody-associated ON (MOG-ON). METHODS: A multicenter validation study was conducted on the published IED cutoff values (>4% or >4 µm in the macular ganglion cell and inner plexiform layer [mGCIP] or >5% or >5 µm in the peripapillary retinal nerve fiber layer [pRNFL]) in individuals with MOG-ON and age-matched and sex-matched healthy controls (HCs). Structural data were acquired with Spectralis spectral-domain OCT >6 months after ON. We calculated sensitivity, specificity, and receiver operating characteristics for both intereye percentage (IEPD) and absolute difference (IEAD). RESULTS: A total of 66 individuals were included (MOG-ON N = 33; HCs N = 33). ON was unilateral in 20 and bilateral in 13 subjects. In the pooled analysis, the mGCIP IEPD was most sensitive (92%), followed by the mGCIP IEAD (88%) and pRNFL (84%). The same pattern was found for the specificity (mGCIP IEPD 82%, IEAD 82%; pRNFL IEPD 82%, IEAD 79%).In subgroup analyses, the diagnostic sensitivity was higher in subjects with unilateral ON (>99% for all metrics) compared with bilateral ON (61%-78%). DISCUSSION: In individuals with MOG-ON, the diagnostic accuracy of OCT-based IED metrics for ON was high, especially of mGCIP IEPD. CLASSIFICATION OF EVIDENCE: This study provides Class III evidence that the intereye difference on OCT can distinguish between those with MOG and normal controls.

A rapid on-site loop-mediated isothermal amplification technology as an early warning system for the detection of Shiga toxin-producing Escherichia coli in water.

Shiga toxin-producing Escherichia coli (STEC) is an important waterborne pathogen capable of causing serious gastrointestinal infections with potentially fatal complications, including haemolytic-uremic syndrome. All STEC serogroups harbour genes that encode at least one Shiga toxin (stx1 and/or stx2), which constitute the primary virulence factors of STEC. Loop-mediated isothermal amplification (LAMP) enables rapid real-time pathogen detection with a high degree of specificity and sensitivity. The aim of this study was to develop and validate an on-site portable diagnostics workstation employing LAMP technology to permit rapid real-time STEC detection in environmental water samples. Water samples (n=28) were collected from groundwater wells (n=13), rivers (n=12), a turlough (n=2) and an agricultural drain (n=1) from the Corrib catchment in Galway. Water samples (100 ml) were passed through a 0.22 µm filter, and buffer was added to elute captured cells. Following filtration, eluates were tested directly using LAMP assays targeting stx1, stx2 and E. coli phoA genes. The portable diagnostics workstation was used in field studies to demonstrate the on-site testing capabilities of the instrument. Real-time PCR assays targeting stx1 and stx2 genes were used to confirm the results. The limit of detection for stx1, stx2 and phoA LAMP assays were 2, 2 and 6 copies, respectively. Overall, stx1, stx2 and phoA genes were detected by LAMP in 15/28 (53.6 %), 9/28 (32.2 %) and 24/28 (85.7 %) samples, respectively. For confirmation, the LAMP results for stx1 and stx2 correlated perfectly (100 %) with those obtained using PCR. The portable diagnostics workstation exhibited high sensitivity throughout the on-site operation, and the average time from sample collection to final result was 40 min. We describe a simple, transferable and efficient diagnostic technology for on-site molecular analysis of various water sources. This method allows on-site testing of drinking water, enabling evidence-based decision-making by public health and water management authorities.

Assessment of hearing dysfunction in patients with Graves' disease and thyroid eye disease without or with teprotumumab.

IMPORTANCE: Thyroid eye disease (TED) negatively impacts quality of life. TED occurs predominantly in Graves' disease (GD). Teprotumumab improves TED but concern for hearing adverse events (AEs) has emerged. Hearing dysfunction is reported in thyroid autoimmune disease but the background prevalence in GD/TED without teprotumumab remains uncertain. OBJECTIVE: To quantify ear-related diagnostic codes/hearing AEs in GD, TED, and patients receiving teprotumumab by examining medical claims and clinical trials. DESIGN AND PARTICIPANTS: Deidentified claims for ear/labyrinth-related ICD-10 codes (KOMODO®) were examined in GD patients without TED, and TED patients without/with teprotumumab treatment. Hearing AE incidence/severity was evaluated in teprotumumab clinical trials. Graves' Ophthalmopathy QOL (GO-QOL) scores were compared in teprotumumab TED trial patients without/with hearing AEs. RESULTS: GD (469,720), TED (38,566) and teprotumumab-treated (967) patients were identified in the claims database. Ear-related codes (including those not specific for hearing) occurred in 24% GD, 33% TED, and 32% teprotumumab-treated patients. "Sensorineural hearing loss bilateral" was most frequent: 32,961/469,720 (7%) GD, 4,279/38,566 (11.1%) TED, and 104/967 (10.8%) teprotumumab patients. Pre-teprotumumab use,165 (17.1%) patients had ear-related codes while 98 (10.1%) had new ear-related codes post-treatment.Eight teprotumumab oncology trials revealed 8.1% (51/633) had Ear/Labyrinth Disorders with 2.1% (13) considered study-drug-related and 3.8% (24) hearing impairment/tinnitus-related AEs, with 1.3% (8) considered teprotumumab-related. Similar rates occurred in TED trials.GO-QOL improved in teprotumumab-treated patients without/with hearing AEs. Incidence/severity was consistent across patients with chronic and acute TED. CONCLUSIONS: These analyses indicate similar occurrence of hearing claims in patients with GD/TED alone as following teprotumumab treatment. Future analyses of incremental hearing risk from teprotumumab should utilize a priori study designs accounting for background hearing dysfunction in patients with GD/TED.

Revisiting the dipeptidyl carboxypeptidase inhibitor captopril as a source of pan anti-trypanosomatid agents.

The protozoan parasites Trypanosoma brucei, Trypanosoma cruzi and Leishmania spp. are responsible for continued propagation of neglected tropical diseases such as African sleeping sickness, Chagas disease and leishmaniasis respectively. Following a report that captopril targets Leishmania donovani dipeptidyl carboxypeptidase, a series of simple proline amides and captopril analogues were synthesized and found to exhibit 1-2 µM in vitro inhibition and selectivity against Trypanosoma brucei, Trypanosoma cruzi and Leishmania spp. The results were corroborated with computational docking studies. Arguably, the synthetic proline amides represent the structurally simplest examples of in vitro pan antiprotozoal compounds.

TSHR-IGF-IR complex drives orbital fibroblast misbehavior in thyroid eye disease.

PURPOSE OF REVIEW: Evolving understanding of thyroid eye disease (TED) has led to rapidly advancing therapeutic options. Most new treatments under development or recently available to patients are predicated on insights into disease mechanism. RECENT FINDINGS: TED, a disfiguring process, involves inflammation and remodeling of the connective tissues around the eye. TED most frequently presents as a component of Graves' disease. Advances in our understanding of cells involved in TED and their molecular interactions have led to novel therapeutic targets. Among these cell types are orbital fibroblasts and a subset comprising monocyte progenitor cells, known as CD34 + CXCR4 + fibrocytes. Among the attributes of fibrocytes is their expression of several autoantigens associated with Graves' disease, including TSHR, thyroglobulin and thyroperoxidase. Fibrocytes also express high levels of the insulin-like growth factor-I (IGF-I) receptor, thought to mediate fibroblast activation. Therapeutically targeting the TSHR/IGF-IR receptor complex using an IGF-I receptor antagonist, teprotumumab, has resulted in substantial clinical benefit for patients with TED. The neural axon repellent, Slit2, and its cognate receptor, ROBO1, appear to modulate the inflammatory phenotype of these orbit-infiltrating fibrocytes. SUMMARY: More detailed understanding of orbital fibroblasts and the distinctions between cell subsets comprising them should lead to more effective therapies with fewer side effects.

Cryo-EM structure of the R388 plasmid conjugative pilus reveals a helical polymer characterized by an unusual pilin/phospholipid binary complex.

Bacterial conjugation is a process by which DNA is transferred unidirectionally from a donor cell to a recipient cell. It is the main means by which antibiotic resistance genes spread among bacterial populations. It is crucially dependent upon the elaboration of an extracellular appendage, termed "pilus," by a large double-membrane-spanning secretion system termed conjugative "type IV secretion system." Here we present the structure of the conjugative pilus encoded by the R388 plasmid. We demonstrate that, as opposed to all conjugative pili produced so far for cryoelectron microscopy (cryo-EM) structure determination, the conjugative pilus encoded by the R388 plasmid is greatly stimulated by the presence of recipient cells. Comparison of its cryo-EM structure with existing conjugative pilus structures highlights a number of important differences between the R388 pilus structure and that of its homologs, the most prominent being the highly distinctive conformation of its bound lipid.

Sterol 14-alpha demethylase (CYP51) activity in Leishmania donovani is likely dependent upon cytochrome P450 reductase 1.

Liposomal amphotericin B is an important frontline drug for the treatment of visceral leishmaniasis, a neglected disease of poverty. The mechanism of action of amphotericin B (AmB) is thought to involve interaction with ergosterol and other ergostane sterols, resulting in disruption of the integrity and key functions of the plasma membrane. Emergence of clinically refractory isolates of Leishmania donovani and L. infantum is an ongoing issue and knowledge of potential resistance mechanisms can help to alleviate this problem. Here we report the characterisation of four independently selected L. donovani clones that are resistant to AmB. Whole genome sequencing revealed that in three of the moderately resistant clones, resistance was due solely to the deletion of a gene encoding C24-sterol methyltransferase (SMT1). The fourth, hyper-resistant resistant clone (>60-fold) was found to have a 24 bp deletion in both alleles of a gene encoding a putative cytochrome P450 reductase (P450R1). Metabolic profiling indicated these parasites were virtually devoid of ergosterol (0.2% versus 18% of total sterols in wild-type) and had a marked accumulation of 14-methylfecosterol (75% versus 0.1% of total sterols in wild-type) and other 14-alpha methylcholestanes. These are substrates for sterol 14-alpha demethylase (CYP51) suggesting that this enzyme may be a bona fide P450R specifically involved in electron transfer from NADPH to CYP51 during catalysis. Deletion of P450R1 in wild-type cells phenocopied the metabolic changes observed in our AmB hyper-resistant clone as well as in CYP51 nulls. Likewise, addition of a wild type P450R1 gene restored sterol profiles to wild type. Our studies indicate that P450R1 is essential for L. donovani amastigote viability, thus loss of this gene is unlikely to be a driver of clinical resistance. Nevertheless, investigating the mechanisms underpinning AmB resistance in these cells provided insights that refine our understanding of the L. donovani sterol biosynthetic pathway.

Retinal Changes in Double-Antibody Seronegative Neuromyelitis Optica Spectrum Disorders.

BACKGROUND AND OBJECTIVES: To systematically describe the clinical picture of double-antibody seronegative neuromyelitis optica spectrum disorders (DN-NMOSD) with specific emphasis on retinal involvement. METHODS: Cross-sectional data of 25 people with DN-NMOSD (48 eyes) with and without a history of optic neuritis (ON) were included in this study along with data from 25 people with aquaporin-4 antibody seropositive neuromyelitis optica spectrum disorder (AQP4-NMOSD, 46 eyes) and from 25 healthy controls (HCs, 49 eyes) for comparison. All groups were matched for age and sex and included from the collaborative retrospective study of retinal optical coherence tomography (OCT) in neuromyelitis optica (CROCTINO). Participants underwent OCT with central postprocessing and local neurologic examination and antibody testing. Retinal neurodegeneration was quantified as peripapillary retinal nerve fiber layer thickness (pRNFL) and combined ganglion cell and inner plexiform layer thickness (GCIPL). RESULTS: This DN-NMOSD cohort had a history of [median (inter-quartile range)] 6 (5; 9) attacks within their 5 ± 4 years since onset. Myelitis and ON were the most common attack types. In DN-NMOSD eyes after ON, pRNFL (p < 0.001) and GCIPL (p = 0.023) were thinner compared with eyes of HCs. Even after only one ON episode, DN-NMOSD eyes already had considerable neuroaxonal loss compared with HCs. In DN-NMOSD eyes without a history of ON, pRNFL (p = 0.027) and GCIPL (p = 0.022) were also reduced compared with eyes of HCs. However, there was no difference in pRNFL and GCIPL between DN-NMOSD and AQP4-NMOSD for the whole group and for subsets with a history of ON and without a history of ON-as well as between variances of retinal layer thicknesses. DISCUSSION: DN-NMOSD is characterized by severe retinal damage after ON and attack-independent retinal neurodegeneration. Most of the damage occurs during the first ON episode, which highlights the need for better diagnostic markers in DN-NMOSD to facilitate an earlier diagnosis as well as for effective and early treatments. In this study, people with DN-NMOSD presented with homogeneous clinical and imaging findings potentially suggesting a common retinal pathology in these patients.

Long-Term Efficacy of Teprotumumab in Thyroid Eye Disease: Follow-Up Outcomes in Three Clinical Trials.

Introduction: Thyroid eye disease (TED) is an autoimmune process characterized by extraocular muscle and orbital fat remodeling/expansion resulting in swelling, pain, redness, proptosis, and diplopia. Teprotumumab, an insulin-like growth factor-I receptor inhibitor, demonstrated improvements in TED signs and symptoms in three adequately powered clinical trials of 24 weeks duration. Here we analyze the long-term maintenance of responses with teprotumumab from these trials. Methods: A total of 112 patients who received 7 or 8 infusions of teprotumumab in the Phase 2, Phase 3 (OPTIC study), and OPTIC Extension (OPTIC-X) studies were included in this analysis. Responses, including clinical activity score (CAS ≥2-point improvement), the European Group of Graves' Orbitopathy ophthalmic composite outcome, diplopia (≥1 Gorman grade improvement), proptosis (≥2 mm improvement), Overall (improvement in proptosis + CAS), and disease inactivation (CAS ≤1), were assessed and pooled from study baseline to week 24 (formal study) and up to week 72 (formal follow-up). Graves' Ophthalmopathy quality-of-life (GO-QoL) scores were also assessed. Outcomes included the percentages of observed patient responses from the study baseline. Additional alternative treatments for TED were assessed as a surrogate of persistent benefit from week 24 through week 120 (extended follow-up). Studies differed in the timing of follow-up visits, and data from some visits were unavailable. Results: At week 72, 52/57 (91.2%), 51/57 (89.5%), 35/48 (72.9%), 38/56 (67.9%), and 37/56 (66.1%) of patients were responders for CAS, composite outcome, diplopia, proptosis, and Overall response, respectively. The mean reduction in proptosis was 2.68 mm (SD 1.92, n = 56), mean GO-QoL improvement was 15.22 (SE 2.82, n = 56), and disease inactivation (CAS ≤1) was detected in 40/57 (70.2%). Over 99 weeks following teprotumumab therapy, 19/106 (17.9%) patients reported additional TED therapy during formal and extended follow-up. Conclusion: The long-term response to teprotumumab as observed 51 weeks after therapy was similar to week 24 results in the controlled clinical trials. Inflammatory and ophthalmic composite outcome improvements were seen in 90% of patients with nearly 70% reporting improvement in diplopia and proptosis. Further, 82% of patients in this analysis did not report additional TED treatment (including surgery) over 99 weeks following the final teprotumumab dose.

Bridging and Validation of the Specific Graves Ophthalmopathy Quality of Life Questionnaire With Health State Utility Values.

OBJECTIVE: In thyroid eye disease (TED), inflammation and expansion of orbital muscle and periorbital fat result in diplopia and proptosis, severely impacting patient quality of life (QOL). The reported health state utility (HSU) scores, which are QOL measures, allow quantification of TED impact and improvement with therapies; however, no current QOL instrument has been validated with HSU scores for TED. Here, we used the disease-specific Graves Ophthalmopathy Quality of Life (GO-QOL) questionnaire and HSU scores to validate QOL impact. METHODS: The GO-QOL scores from patients in 2 randomized, masked, placebo-controlled teprotumumab trials (N=171) were compared with 6 HSU values based on severity of proptosis/diplopia in those studies. Patient GO-QOL and HSU scores were compared at baseline and after 6-month treatment via regression analyses. GO-QOL and HSU scores were correlated for validation and quantification of QOL impact by severity state and to estimate quality-adjusted life year improvement. RESULTS: GO-QOL scores were correlated with TED severity, indicating that worse severity was associated with lower (worse) GO-QOL scores. Less severe health states were represented by higher (better) GO-QOL scores. Importantly, GO-QOL scores were positively correlated with utility scores of the 6 health states, allowing for conversion of the GO-QOL scores to utility scores. A positive (improved) 0.013 utility change was found for each 1-point (positive) improvement in GO-QOL score produced by teprotumumab versus placebo. CONCLUSION: Patients with moderate-to-severe active TED health states demonstrate increasing TED severity associated with declining utility values and worsening GO-QOL scores. These results indicate that the GO-QOL scores can be used to bridge to the HSU scores for benefit quantification.

Glycemic Trends in Patients with Thyroid Eye Disease Treated with Teprotumumab in 3 Clinical Trials.

PURPOSE: Assess incidence, severity, and glucose excursion outcomes in thyroid eye disease (TED) patients receiving the insulin-like growth factor-1 receptor inhibitor teprotumumab from 3 clinical trials. DESIGN: Analysis of pooled glycemic data over time. PARTICIPANTS: Eighty-four teprotumumab- and 86 placebo-treated active TED patients from the phase 2 and phase 3 (OPTIC) controlled clinical trials and 51 teprotumumab-treated patients from the OPTIC extension (OPTIC-X) trial. METHODS: Eight intravenous infusions were given over 21 weeks. Phase 2 serum glucose was measured at weeks 1, 4, 15, and 21, with fasting measurements at weeks 1 and 4. Serum glucose was measured at each study visit in OPTIC and OPTIC-X, with fasting measurements at weeks 1 and 4 (in patients without diabetes) or all visits (in patients with diabetes). In all studies, hemoglobin A1c (HbA1c) was measured at baseline, 12, and 24 weeks plus weeks 36 and 48 in OPTIC-X. MAIN OUTCOME MEASURES: Serum glucose and HbA1c. RESULTS: In the phase 2 and 3 studies, 9 hyperglycemic episodes occurred in 8 teprotumumab patients; mean HbA1c level increased 0.22% from baseline to week 24 (to 5.8%; range, 5.0%-7.9%) versus 0.04% in patients receiving the placebo (to 5.6%; range, 4.6%-8.1%). At study end, 78% (59/76) of teprotumumab patients and 87% (67/77) of patients receiving placebo had normoglycemic findings. Normoglycemia was maintained in 84% (57/68) of patients receiving teprotumumab and 93% (64/69) of patients receiving placebo. Among baseline prediabetic patients, 43% (3/7) remained prediabetic in both groups, and 29% (2/7) of teprotumumab patients and 14% (1/7) of patients receiving placebo had diabetic findings at week 24. OPTIC-X patients trended toward increased fasting glucose and HbA1c whether initially treated or retreated with teprotumumab. Fasting glucose commonly rose after 2 or 3 infusions and stabilized thereafter. Most hyperglycemic incidents occurred in patients with baseline prediabetes/diabetes but were controlled with medication. No evidence was found for progression or increased incidence of hyperglycemia with subsequent doses. CONCLUSIONS: Serious glycemic excursions are uncommon in patients with normoglycemia before teprotumumab therapy. Patients with controlled diabetes or impaired glucose tolerance can be treated safely if baseline screening, regular monitoring of glycemic control, and timely treatment of hyperglycemia are practiced. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

The role of ion dissolution in metal and metal oxide surface inactivation of SARS-CoV-2.

Anti-viral surface coatings are under development to prevent viral fomite transmission from high-traffic touch surfaces in public spaces. Copper's anti-viral properties have been widely documented, but the anti-viral mechanism of copper surfaces is not fully understood. We screened a series of metal and metal oxide surfaces for anti-viral activity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease (COVID-19). Copper and copper oxide surfaces exhibited superior anti-SARS-CoV-2 activity; however, the level of anti-viral activity was dependent on the composition of the carrier solution used to deliver virus inoculum. We demonstrate that copper ions released into solution from test surfaces can mediate virus inactivation, indicating a copper ion dissolution-dependent anti-viral mechanism. The level of anti-viral activity is, however, not dependent on the amount of copper ions released into solution per se. Instead, our findings suggest that degree of virus inactivation is dependent on copper ion complexation with other biomolecules (e.g., proteins/metabolites) in the virus carrier solution that compete with viral components. Although using tissue culture-derived virus inoculum is experimentally convenient to evaluate the anti-viral activity of copper-derived test surfaces, we propose that the high organic content of tissue culture medium reduces the availability of "uncomplexed" copper ions to interact with the virus, negatively affecting virus inactivation and hence surface anti-viral performance. We propose that laboratory anti-viral surface testing should include virus delivered in a physiologically relevant carrier solution (saliva or nasal secretions when testing respiratory viruses) to accurately predict real-life surface anti-viral performance when deployed in public spaces.IMPORTANCEThe purpose of evaluating the anti-viral activity of test surfaces in the laboratory is to identify surfaces that will perform efficiently in preventing fomite transmission when deployed on high-traffic touch surfaces in public spaces. The conventional method in laboratory testing is to use tissue culture-derived virus inoculum; however, this study demonstrates that anti-viral performance of test copper-containing surfaces is dependent on the composition of the carrier solution in which the virus inoculum is delivered to test surfaces. Therefore, we recommend that laboratory surface testing should include virus delivered in a physiologically relevant carrier solution to accurately predict real-life test surface performance in public spaces. Understanding the mechanism of virus inactivation is key to future rational design of improved anti-viral surfaces. Here, we demonstrate that release of copper ions from copper surfaces into small liquid droplets containing SARS-CoV-2 is a mechanism by which the virus that causes COVID-19 can be inactivated.

Real-time single-base specific detection of the Haemonchus contortus S168T variant associated with levamisole resistance using loop-primer endonuclease cleavage loop-mediated isothermal amplification.

Haemonchus contortus is a parasitic haematophagous nematode that primarily affects small ruminants and causes significant economic loss to the global livestock industry. Treatment of haemonchosis typically relies on broad-spectrum anthelmintics, resistance to which is an important cause of treatment failure. Resistance to levamisole remains less widespread than to other major anthelmintic classes, prompting the need for more effective and accurate surveillance to maintain its efficacy. Loop-primer endonuclease cleavage loop-mediated isothermal amplification (LEC-LAMP) is a recently developed diagnostic method that facilitates multiplex target detection with single nucleotide polymorphism (SNP) specificity and portable onsite testing. In this study, we designed a new LEC-LAMP assay and applied it to detect the levamisole resistance marker S168T in H. contortus. We explored multiplexing probes for both the resistant S168T and the susceptible S168 alleles in a single-tube assay. We then included a generic probe to detect the acr-8 gene in the multiplex assay, which could facilitate the quantification of both resistance markers and overall genetic material from H. contortus in a single step. Our results showed promising application of these technologies, demonstrating a proof-of-concept assay which is amenable to detection of resistance alleles within the parasite population, with the potential for multiplex detection, and point-of-care application enabled by lateral flow end-point detection. However, further optimisation and validation is necessary.

Fibrocyte Participation in Thyroid-Associated Ophthalmopathy Suggests New Approaches to Therapy.

PURPOSE: Review the historical context of research and changing therapeutic landscape of thyroid-associated ophthalmopathy (TAO) by focusing on the relationship between TAO, CD34+ fibrocytes, thyrotropin receptor (TSHR), and insulin-like growth factor-I receptor (IGF-IR). METHODS: A literature review using search terms, including fibrocytes, IGF-IR, TSHR, TAO, and thyroid eye disease. RESULTS: The mechanisms involved in TAO have been partially identified. Substantial progress has been made over several decades, including 1) recognizing the interplay between the professional immune system and orbital tissues; 2) TSHR and IGF-IR act interdependently in mediating the pathogenesis of TAO; 3) Multiple cytokines and specific immune cells are involved in activating and remodeling orbital tissue; 4) Recognition of these mechanisms is allowing the development of target therapies such as teprotumumab, a monoclonal antibody IGF-IR inhibitor approved by the US Food and drug administration for treatment of TAO; and 5) It appears that teprotumumab acts on the systemic immune system peripheral to the orbit. CONCLUSION: Additional molecules targeting IGF-IR and other plausible disease mechanisms are currently under development. This activity in the TAO therapeutic space portends even greater improvements in patient care.

A type VII-secreted lipase toxin with reverse domain arrangement.

The type VII protein secretion system (T7SS) is found in many Gram-positive bacteria and in pathogenic mycobacteria. All T7SS substrate proteins described to date share a common helical domain architecture at the N-terminus that typically interacts with other helical partner proteins, forming a composite signal sequence for targeting to the T7SS. The C-terminal domains are functionally diverse and in Gram-positive bacteria such as Staphylococcus aureus often specify toxic anti-bacterial activity. Here we describe the first example of a class of T7 substrate, TslA, that has a reverse domain organisation. TslA is widely found across Bacillota including Staphylococcus, Enterococcus and Listeria. We show that the S. aureus TslA N-terminal domain is a phospholipase A with anti-staphylococcal activity that is neutralised by the immunity lipoprotein TilA. Two small helical partner proteins, TlaA1 and TlaA2 are essential for T7-dependent secretion of TslA and at least one of these interacts with the TslA C-terminal domain to form a helical stack. Cryo-EM analysis of purified TslA complexes indicate that they share structural similarity with canonical T7 substrates. Our findings suggest that the T7SS has the capacity to recognise a secretion signal present at either end of a substrate.

Patient Perspectives on a Tailored Self-Management Education and Support Intervention for Low-Income Seniors With Chronic Health Conditions.

Background: The Assessing Outcomes of Enhanced Chronic Disease Care Through Patient Education and a Value-based Formulary Study (ACCESS) was a 2 x 2 factorial randomized trial that tested the impact of a tailored self-management education support (SMES) program, which demonstrated a 22% reduction in adverse clinical events. We sought to qualitatively explore participants' perspectives on the SMES intervention, and the ways in which it may have improved self-management skills. Methods: We used a qualitative descriptive approach and conducted individual semistructured interviews. We conducted inductive and deductive thematic analysis using NVivo 12 (QSR International, Burlington, MA). Results: We interviewed 20 participants who had recently completed the 3-year SMES intervention. The following 3 main themes emerged from the data: (i) empowerment; (ii) intervention acceptability; and (iii) suggestions for improvement. Regarding empowerment, we identified subthemes of health literacy, self-efficacy, self-management, and active role in health. Several participants reported that empowerment promoted health behaviour change or improved confidence in self-management. Regarding acceptability, we identified subthemes of ease of use and presentation style. Most participants expressed positive feelings toward the intervention and felt that it was easy to understand. Finally, we identified subthemes of learning style, content, and engagement strategies, within the theme of suggestions for improvement. Some participants said that the messages were too general and did not fully address the complex health concerns they had. Conclusions: Our results highlighted key strategies to promote patient engagement and self-management behaviours and demonstrated how they may have been used to improve clinical endpoints. Additionally, we demonstrated the novel use of marketing principles in SMES interventions.

Contexte: L'étude ACCESS (pour Assessing Outcomes of Enhanced Chronic Disease Care Through Patient Education and a Value-based Formulary Study) était un essai à répartition aléatoire avec un plan factoriel 2 x 2 qui a mesuré l'effet d'un programme personnalisé de soutien à la formation sur l'autogestion dans laquelle une réduction de 22 % des événements cliniques défavorables a été observée. Notre objectif était de réaliser une exploration qualitative du point de vue des patients au sujet de l'intervention et des façons dont elle a permis d'améliorer leurs habiletés d'autogestion. Méthodologie: Nous avons utilisé une approche descriptive et qualitative et nous avons mené des entretiens individuels semi-structurés auprès des participants. Des analyses thématiques inductive et déductive ont été réalisées avec NVivo 12 (QSR International, Burlington MA). Résultats: Des entretiens ont été menés auprès de 20 personnes ayant récemment terminé l'intervention de 3 ans. Les données recueillies ont permis de cerner 3 thèmes principaux : (i) l'autonomisation; (ii) l'acceptabilité de l'intervention; et (iii) les suggestions pour l'amélioration du programme. En ce qui concerne l'autonomisation des patients, nous avons relevé les sous-thèmes de la littératie dans le domaine de la santé, de l'auto-efficacité, de l'autogestion et de la participation active dans le domaine de la santé. Plusieurs participants ont mentionné que l'autonomisation avait favorisé des changements de comportements liés à la santé ou avait amélioré leur niveau de confiance quant à leur autogestion. Pour ce qui est de l'acceptabilité, nous avons noté les sous-thèmes de la facilité d'utilisation et du style des présentations. La plupart des participants ont exprimé une opinion favorable au sujet de l'intervention et la trouvaient facile à comprendre. En dernier lieu, nous avons relevé les thèmes des styles d'apprentissage, du contenu et des stratégies de mobilisation, que nous avons regroupés sous le thème des suggestions d'amélioration. Certains participants ont mentionné que les messages étaient trop généraux et n'abordaient pas leurs préoccupations complexes liées à la santé. Conclusions: Les résultats que nous avons obtenus ont mis en évidence des stratégies clés pour favoriser la participation des patients et leurs comportements d'autogestion et la façon dont elles ont pu améliorer les résultats cliniques de patients. De plus, nous avons démontré une nouvelle utilisation de principes tirés du domaine du marketing dans des interventions de soutien à la formation sur l'autogestion.