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Abstract Background Internal control genes with highly uniform expression throughout the experimental conditions are required for accurate gene expression analysis as no universal reference genes exists. In this study, the expression stability of 24 candidate genes from Triticum aestivum cv. Cubus flag leaves grown under organic and conventional farming systems was evaluated in two locations in order to select suitable genes that can be used for normalization of real-time quantitative reverse-transcription PCR (RT-qPCR) reactions. The genes were selected among the most common used reference genes as well as genes encoding proteins involved in several metabolic pathways. Findings Individual genes displayed different expression rates across all samples assayed. Applying geNorm, a set of three potential reference genes were suitable for normalization of RT-qPCR reactions in winter wheat flag leaves cv. Cubus: TaFNRII (ferredoxin-NADP(H) oxidoreductase; AJ457980.1), ACT2 (actin 2; TC234027), and rrn26 (a putative homologue to RNA 26S gene; AL827977.1). In addition of these three genes that were also top-ranked by NormFinder, two extra genes: CYP18-2 (Cyclophilin A, AY456122.1) and TaWIN1 (14-3-3 like protein, AB042193) were most consistently stably expressed. Furthermore, we showed that TaFNRII, ACT2, and CYP18-2 are suitable for gene expression normalization in other two winter wheat varieties (Tommi and Centenaire) grown under three treatments (organic, conventional and no nitrogen) and a different environment than the one tested with cv. Cubus. Conclusions This study provides a new set of reference genes which should improve the accuracy of gene expression analyses when using wheat flag leaves as those related to the improvement of nitrogen use efficiency for cereal production.

Otitis externa is an inflammatory disease of the external ear canal of complex and multifactorial etiology associated with recurrent bacterial infection. This study aimed to assess the antimicrobial and antibiofilm activity of promethazine against bacterial isolates from dogs with otitis externa, as well as the effect of this compound on the dynamics of biofilm formation over 120 h. Planktonic bacterial susceptibility to promethazine was evaluated to determine the minimum inhibitory concentrations (MIC). The minimum biofilm eradication concentration (MBEC) was also determined by broth microdilution. To evaluate the effect on biofilm growth, promethazine was tested at three concentrations MIC, MIC/2 and MIC/8, with daily readings at 48, 72, 96 and 120 h. The MICs of promethazine ranged from 48.83 to 781.25 μg mL -1 . Promethazine significantly (P < 0.05) reduced mature biofilm biomass, with MBECs ranging from 48.8 to 6250 μg mL -1 and reduced (P < 0.01) biofilm formation for up to the 120-h, at concentrations corresponding to the MIC obtained against each isolate. Promethazine was effective against microorganisms associated with canine otitis externa. The data suggest that promethazine presents antimicrobial and antibiofilm activity and is a potential alternative to treat and prevent recurrent bacterial otitis in dogs. These results emphasize the importance of drug repurposing in veterinary otology as an alternative to reduce antimicrobial resistance., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Debora Castelo-Branco reports financial support was provided by National Council for Scientific and Technological Development. Jose Sidrim reports financial support was provided by National Council for Scientific and Technological Development. None to declare. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Candida albicans invasive candidiasis is considered a global health problem. In such cases, biofilm formation on implanted devices represents a therapeutic challenge and the presence of metabolically inactive persistent cells (PCs) in these communities increases their tolerance to fungicidal drugs. This study investigated the influence of amoxicillin, AMX; cefepime, CEF; gentamicin, GEN; amikacin, AMK; vancomycin, VAN; and ciprofloxacin, CIP; on the production of PCs in biofilms of C. albicans bloodstream isolates. 48 h-mature biofilms ( n  = 6) grown in RPMI-1640 supplemented with antibiotics were treated with 100 μg ml -1 amphotericin B and then evaluated for PCs. Biofilms grown in the presence of antibiotics produced more PCs, up to 10×, when exposed to AMX and CIP; 5 × to CEF; and 6 × to GEN and VAN. The results indicate that antibiotics can modulate PC production in C. albicans biofilms. This scenario may have clinical repercussions in immunocompromised patients under broad-spectrum antibiotic therapy.

Although the terms "rare diseases" (RD) and "orphan diseases" (OD) are often used interchangeably, specific nuances in definitions should be noted to avoid misconception. RD are characterized by a low prevalence within the population, whereas OD are those inadequately recognized or even neglected by the medical community and drug companies. Despite their rarity, as our ability on discovering novel clinical phenotypes and improving diagnostic tools expand, RD will continue posing a real challenge for rheumatologists. Over the last decade, there has been a growing interest on elucidating mechanisms of rare autoimmune and autoinflammatory rheumatic diseases, allowing a better understanding of the role played by immune dysregulation on granulomatous, histiocytic, and hypereosinophilic disorders, just to name a few. This initiative enabled the rise of innovative targeted therapies for rheumatic RD. In this review, we explore the state-of-the art of rare RD and the critical role played by rheumatologists in healthcare. We also describe the challenges rheumatologists may face in the coming decades., (© 2024. The Author(s).)

Amyloidosis is a localized or systemic disease caused by deposition of proteins in the extracellular space of various organs and tissues. As part of the disease, proteins that were originally soluble misfold and acquire a fibrillar conformation that renders them insoluble and resistant to proteolysis. Systemic amyloidosis is a rare, often underdiagnosed condition. In recent years, the incidence of newly diagnosed cases of amyloidosis has been increasing in association with the aging of the population and greater access to diagnostic tests. From a clinical perspective, systemic amyloidosis is frequently associated with involvement of the kidneys (causing nephrotic syndrome), heart (cardiac failure and arrhythmia), and peripheral nervous system (sensorimotor polyneuropathy and autonomic dysfunction). This condition is important to the rheumatologist for several reasons, such as its systemic involvement that mimics autoimmune rheumatic diseases, its musculoskeletal manifestations, which when recognized can allow the diagnosis of amyloidosis, and also because reactive or secondary AA amyloidosis is a complication of rheumatic inflammatory diseases. The treatment of amyloidosis depends on the type of amyloid protein involved. Early recognition of this rare disease is fundamental for improved clinical outcomes., (© 2024. The Author(s).)

Efflux pump inhibitors are a potential therapeutic strategy for managing antimicrobial resistance and biofilm formation. This article evaluated the effect of carbonyl cyanide m-chlorophenyl hydrazone (CCCP) on the biofilm growth dynamics and the production of virulence factors by Burkholderia pseudomallei . The effects of CCCP on planktonic, growing, and mature biofilm, interaction with antibacterial drugs, and protease and siderophore production were assessed. CCCP MICs ranged between 128 and 256 µM. The CCCP (128 µM) had a synergic effect with all the antibiotics tested against biofilms. Additionally, CCCP reduced ( p  < .05) the biomass of biofilm growth and mature biofilms at 128 and 512 µM, respectively. CCCP also decreased ( p  < .05) protease production by growing (128 µM) and induced ( p  < .05) siderophore release by planktonic cells (128 µM) growing biofilms (12.8 and 128 µM) and mature biofilms (512 µM). CCCP demonstrates potential as a therapeutic adjuvant for disassembling B. pseudomallei biofilms and enhancing drug penetration.

Primary Sjögren's syndrome (pSS) is a systemic autoimmune disease that affects exocrine glands, mainly the salivary and lacrimal glands, leading to the development of sicca symptoms. Patients with pSS may also present with extraglandular manifestations, including lung involvement, estimated to occur in 9-24% of cases. Shrinking lung syndrome (SLS) is an uncommon respiratory complication primarily associated with systemic lupus erythematosus, with a prevalence of approximately 1% in these patients. It typically manifests as dyspnea, pleuritic chest pain, lung volume reduction, and a restrictive pattern on respiratory function tests. Cases reporting SLS with other connective tissue diseases, including pSS, are even rarer. Herein, we describe a case of a 57-year-old woman with a 10-year history of pSS who presented with dyspnea and pleuritic chest pain. After evaluation, the patient was diagnosed with SLS based on clinical, radiologic, laboratorial, and electrophysiologic characteristics. In addition, we identified and analyzed previously published cases of SLS in pSS. Treatment includes corticosteroids, immunosuppressants, and respiratory muscle training. This study highlights the importance of considering SLS in the differential diagnosis of patients with pSS and respiratory symptoms., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Systemic autoinflammatory diseases (SAIDs) arise from dysregulated innate immune system activity, which leads to systemic inflammation. These disorders, encompassing a diverse array of genetic defects classified as inborn errors of immunity, are significant diagnostic challenges due to their genetic heterogeneity and varied clinical presentations. Although recent advances in genetic sequencing have facilitated pathogenic gene discovery, approximately 40% of SAIDs patients lack molecular diagnoses. SAIDs have distinct clinical phenotypes, and targeted therapeutic approaches are needed. This review aims to underscore the complexity and clinical significance of SAIDs, focusing on prototypical disorders grouped according to their pathophysiology as follows: (i) inflammasomopathies, characterized by excessive activation of inflammasomes, which induces notable IL-1β release; (ii) relopathies, which are monogenic disorders characterized by dysregulation within the NF-κB signaling pathway; (iii) IL-18/IL-36 signaling pathway defect-induced SAIDs, autoinflammatory conditions defined by a dysregulated balance of IL-18/IL-36 cytokine signaling, leading to uncontrolled inflammation and tissue damage, mainly in the skin; (iv) type I interferonopathies, a diverse group of disorders characterized by uncontrolled production of type I interferons (IFNs), notably interferon α, β, and ε; (v) anti-inflammatory signaling pathway impairment-induced SAIDs, a spectrum of conditions characterized by IL-10 and TGFβ anti-inflammatory pathway disruption; and (vi) miscellaneous and polygenic SAIDs. The latter group includes VEXAS syndrome, chronic recurrent multifocal osteomyelitis/chronic nonbacterial osteomyelitis, Schnitzler syndrome, and Still's disease, among others, illustrating the heterogeneity of SAIDs and the difficulty in creating a comprehensive classification. Therapeutic strategies involving targeted agents, such as JAK inhibitors, IL-1 blockers, and TNF inhibitors, are tailored to the specific disease phenotypes., (© 2024. The Author(s).)

Immunoglobulin G4-related disease is a systemic immune-mediated disease with insidious evolution characterized by fibroinflammatory lesions over virtually any organ system. Despite the remarkable progression of knowledge, its etiology remains undefined. Due to its relapse-remitting pattern, it could accumulate irreversible damage, increasing comorbidities and mortality. This paper emphasizes key concepts for diagnosing and treating patients with this condition., (© 2024. The Author(s).)

Background: Fungal keratitis is a severe eye infection that can result in blindness and visual impairment, particularly in developing countries. Fusarium spp. are the primary causative agents of this condition. Diagnosis of Fusarium keratitis (FK) is challenging, and delayed treatment can lead to serious complications. However, there is limited epidemiological data on FK, especially in tropical areas., Objectives: This study aimed to describe the clinical, laboratorial and epidemiological characteristics of FK in a tropical semi-arid region of Brazil., Patients/methods: Adult patients with laboratory-confirmed FK diagnosed between October 2019 and March 2022 were evaluated. Fusarium isolates were characterized at molecular level and evaluated regarding antifungal susceptibility., Results: A total of 226 clinical samples from patients suspected of keratitis were evaluated; fungal growth was detected in 50 samples (22.12%); out of which 42 were suggestive of Fusarium spp. (84%). Molecular analysis of a randomly selected set of 27 isolates identified F. solani species complex (n = 14); F. fujikuroi sensu lato (n = 6) and F. dimerum sensu lato (n = 7); a total of 10 haplotypes were identified among the strains. All but one Fusarium strains were inhibited by amphotericin B, natamycin and fluconazole. Most patients were male (71.42%; 30 out of 42), aged from 27 to 73 years old. Trauma was the most important risk factor for FK (40.47%; 17 out of 42). Patients were treated with antifungals, corticoids and antibiotics; keratoplasty and eye enucleation were also performed., Conclusions: The study provided insights into the characteristics of FK in tropical regions and emphasized the importance of enhanced surveillance and management strategies., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

Gaucher and Fabry diseases are lysosomal storage disorders in which deficient enzyme activity leads to pathological accumulation of sphingolipids. These diseases have a broad phenotypic presentation. Musculoskeletal symptoms and pain complaints are frequently reported by patients. Thus, rheumatologists can be contacted by these patients, contributing to the correct diagnosis, earlier indication of appropriate treatment and improvement of their prognosis. This review describes important concepts about Gaucher and Fabry diseases that rheumatologists should understand to improve patients' quality of life and change the natural history of these diseases., (© 2024. The Author(s).)

Despite their rarity, Lyme disease and Whipple's disease are of significant importance in rheumatology, as both can manifest as chronic arthritis, presenting challenges in the differential diagnosis of inflammatory arthropathies. In Lyme disease, arthritis typically emerges as a late manifestation, usually occurring six months after the onset of erythema migrans. The predominant presentation involves mono- or oligoarthritis of large joints, with a chronic or remitting-recurrent course. Even with appropriate antimicrobial treatment, arthritis may persist due to inadequate immunological control triggered by the disease. In contrast, Whipple's disease may present with a migratory and intermittent seronegative poly- or oligoarthritis of large joints, preceding classic gastrointestinal symptoms by several years. Both disorders, particularly Whipple's disease, can be misdiagnosed as more common autoimmune rheumatic conditions such as rheumatoid arthritis and spondyloarthritis. Epidemiology is crucial in suspecting and diagnosing Lyme disease, as the condition is transmitted by ticks prevalent in specific areas of the United States, Europe, and Asia. On the contrary, the causative agent of Whipple's disease is widespread in the environment, yet invasive disease is rare and likely dependent on host genetic factors. In addition to erythema migrans in Lyme disease and gastrointestinal manifestations in Whipple's disease, neurological and cardiac involvement can further complicate the course of both. This article offers a comprehensive review of the epidemiological, pathophysiological, clinical, and therapeutic aspects of both diseases., (© 2024. The Author(s).)

Aim: Synthetic antimicrobial peptides (SAMPs) present the potential to fight systemic fungal infections. Here, the PHO36 receptor from Candida albicans was analyzed by in silico tools as a possible target for three anticandidal SAMPs: Rc Alb-PepIII, PepGAT and PepKAA. Materials & methods: Molecular docking, dynamics and quantum biochemistry were employed to understand the individual contribution of amino acid residues in the interaction region. Results: The results revealed that SAMPs strongly interact with the PHO36 by multiple high-energy interactions. This is the first study to employ quantum biochemistry to describe the interactions between SAMPs and the PHO36 receptor. Conclusion: This work contributes to understanding and identifying new molecular targets with medical importance that could be used to discover new drugs against systemic fungal infections.

Hypertrophic pachymeningitis (HP) is a rare and chronic inflammatory disorder presenting as localized or diffuse thickening of the dura mater. It can be idiopathic or an unusual manifestation of immune-mediated, infectious, and neoplastic conditions. Although some cases may remain asymptomatic, HP can lead to progressive headaches, cranial nerve palsies, hydrocephalus, and other neurological complications, which makes its recognition a fundamental step for prompt treatment. Regarding the diagnosis workup, enhanced MRI is the most useful imaging method to evaluate dural thickening. This article addresses the MR imaging patterns of immune-mediated HP, including immunoglobulin G4-related disease, neurosarcoidosis, granulomatosis with polyangiitis, rheumatoid pachymeningitis, and idiopathic HP. The main infectious and neoplastic mimicking entities are also discussed with reference to conventional and advanced MR sequences., Competing Interests: Declaration of Competing Interest None., (Copyright © 2023 The Association of University Radiologists. Published by Elsevier Inc. All rights reserved.)

This study evaluated the antibiofilm activity of promethazine, deferiprone, and Manuka honey against Staphylococcus aureus and Pseudomonas aeruginosa in vitro and ex vivo in a wound model on porcine skin. The minimum inhibitory concentrations (MICs) and the effects of the compounds on biofilms were evaluated. Then, counting colony-forming units (CFUs) and confocal microscopy were performed on biofilms cultivated on porcine skin for evaluation of the compounds. For promethazine, MICs ranging from 97.66 to 781.25 µg/ml and minimum biofilm eradication concentration (MBEC) values ranging from 195.31 to 1562.5 µg/ml were found. In addition to reducing the biomass of both species' biofilms. As for deferiprone, the MICs were 512 and >1024 µg/ml, the MBECs were ≥1024 µg/ml, and it reduced the biomass of biofilms. Manuka honey had MICs of 10%-40%, MBECs of 20 to >40% and reduced the biomass of S. aureus biofilms only. Concerning the analyses in the ex vivo model, the compounds reduced (P < .05) CFU counts for both bacterial species, altering the biofilm architecture. The action of the compounds on biofilms in in vitro and ex vivo tests raises the possibility of using them against biofilm-associated wounds. However, further studies are needed to characterize the mechanisms of action and their effectiveness on biofilms in vivo., (© The Author(s) 2023. Published by Oxford University Press on behalf of Applied Microbiology International.)

Demyelinating and inflammatory myelopathies represent a group of diseases with characteristic patterns in neuroimaging and several differential diagnoses. The main imaging patterns of demyelinating myelopathies (multiple sclerosis, neuromyelitis optica spectrum disorder, acute disseminated encephalomyelitis, and myelin oligodendrocyte glycoprotein antibody-related disorder) and inflammatory myelopathies (systemic lupus erythematosus-myelitis, sarcoidosis-myelitis, Sjögren-myelitis, and Behçet's-myelitis) will be discussed in this article, highlighting key points to the differential diagnosis., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Aim: This study evaluated the effect of fluoxetine (FLU) on planktonic and biofilm growth and the antimicrobial susceptibility of Burkholderia pseudomallei . Materials & methods: The minimum inhibitory concentrations (MICs) for FLU were determined by broth microdilution. Its effect on growing and mature biofilms and its interaction with antibacterial drugs were evaluated by assessing biofilm metabolic activity, biomass and structure through confocal microscopy. Results: The FLU MIC range was 19.53-312.5 μg/ml. FLU eradicated growing and mature biofilms of B. pseudomallei at 19.53-312.5 μg/ml and 1250-2500 μg/ml, respectively, with no structural alterations and enhanced the antibiofilm activity of antimicrobial drugs. Conclusion: These results bring perspectives for the use of FLU in the treatment of melioidosis, requiring further studies to evaluate its applicability.

The present study aimed to: (1) evaluate the influence of the steroid hormones (SH) on biofilm development; (2) investigate the formation of persister cells (PC) in biofilms; and (3) investigate the influence of SH on PC formation. Biofilms were derived from vulvovaginal candidiasis (VVC) samples and evaluated by three models: microcosm biofilms grown in Vaginal Fluid Simulator Medium (MiB-VFSM); monospecies biofilms grown in VFSM (MoB-VFSM) and RPMI media (MoB-RPMI). SH altered cell counting and biomass of biofilms grown in VSFM; MoB-RPMI were negatively affected by SH. SH stimulated the formation of PC in MiB-VFSM but not MoB-VFSM; MoB-RPMI showed a lower number of PC in the presence of SH. The results showed that SH altered the dynamics of biofilm formation and development, depending on the study model. The data suggest the influence of hormones on the physiology of Candida biofilms and reinforce the importance of PC in the pathogenesis of VVC.

Systemic autoimmune myopathies (SAMs) are rare diseases that lead to muscle inflammation and may be associated with a variety of systemic manifestations. Although there is great heterogeneity in the spectrum of extra-muscular involvement in SAMs, interstitial lung disease (ILD) is the most frequent lung manifestation. SAM-related ILD (SAM-ILD) presents significant variations according to geographic location and temporal trends and is associated with increased morbidity and mortality. Several myositis autoantibodies have been discovered over the last decades, including antibodies targeting aminoacyl-tRNA synthetase enzymes, which are associated with a variable risk of developing ILD and a myriad of other clinical features. In this review, the most relevant topics regarding clinical manifestations, risk factors, diagnostic tests, autoantibodies, treatment, and prognosis of SAM-ILD are highlighted. We searched PubMed for relevant articles published in English, Portuguese, or Spanish from January 2002 to September 2022. The most common SAM-ILD patterns are nonspecific interstitial pneumonia and organizing pneumonia. The combination of clinical, functional, laboratory, and tomographic features is usually sufficient for diagnostic confirmation, without the need for additional invasive methods. Glucocorticoids remain the first-line treatment for SAM-ILD, although other traditional immunosuppressants, such as azathioprine, mycophenolate, and cyclophosphamide have demonstrated some efficacy and, therefore, have an important role as steroid-sparing agents.

Ex vivo experiments have been performed aiming at mimicking in vivo environments. The main aim of this research was to standardize in vitro dual-species biofilm formation by Staphylococcus pseudintermedius and Malassezia pachydermatis as a strategy to establish an ex vivo biofilm model. Initially, the in vitro formation of biofilms in co-culture was established, using YPD medium, inoculum turbidity of 0.5 on the McFarland scale and maturation periods of 96 h for M. pachydermatis and 48 h for S. pseudintermedius. Subsequently, biofilms were formed on porcine skin using the same conditions, under which a greater number of cells/ml was observed in in vitro dual-species than in in vitro mono-species biofilms. Furthermore, ex vivo biofilm images demonstrated the formation of a highly structured biofilm with the presence of cocci and yeasts surrounded by the matrix. Thus, these conditions optimized the growth of both microorganisms within biofilms in vitro and ex vivo., Competing Interests: Declaration of Competing Interest We declare no conflicts of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Objectives: To compare pain intensity among individuals with idiopathic inflammatory myopathies (IIMs), other systemic autoimmune rheumatic diseases (AIRDs), and without rheumatic disease (wAIDs)., Methods: Data were collected from the COVID-19 Vaccination in Autoimmune Diseases (COVAD) study, an international cross-sectional online survey, from December 2020 to August 2021. Pain experienced in the preceding week was assessed using numeral rating scale (NRS). We performed a negative binomial regression analysis to assess pain in IIMs subtypes and whether demographics, disease activity, general health status, and physical function had an impact on pain scores., Results: Of 6988 participants included, 15.1% had IIMs, 27.9% had other AIRDs, and 57.0% were wAIDs. The median pain NRS in patients with IIMs, other AIRDs, and wAIDs were 2.0 (interquartile range [IQR] = 1.0-5.0), 3.0 (IQR = 1.0-6.0), and 1.0 (IQR = 0-2.0), respectively (P < 0.001). Regression analysis adjusted for gender, age, and ethnicity revealed that overlap myositis and antisynthetase syndrome had the highest pain (NRS = 4.0, 95% CI = 3.5-4.5, and NRS = 3.6, 95% CI = 3.1-4.1, respectively). An additional association between pain and poor functional status was observed in all groups. Female gender was associated with higher pain scores in almost all scenarios. Increasing age was associated with higher pain NRS scores in some scenarios of disease activity, and Asian and Hispanic ethnicities had reduced pain scores in some functional status scenarios., Conclusion: Patients with IIMs reported higher pain levels than wAIDs, but less than patients with other AIRDs. Pain is a disabling manifestation of IIMs and is associated with a poor functional status., (© 2023 The Authors. International Journal of Rheumatic Diseases published by Asia Pacific League of Associations for Rheumatology and John Wiley & Sons Australia, Ltd.)

Bats (Chiroptera) are flying mammals of great biodiversity and habits. These characteristics contribute for them being natural reservoirs and part of the epidemiological cycle of several potentially zoonotic pathogens, such as viruses, protozoa, fungi and bacteria. Brazil hosts approximately 15% of the world's bat diversity, with 181 distinct species, 68 genera and 9 families. About 60% of infectious diseases in humans are of zoonotic origin and, in the last decades, the detection of zoonotic pathogens in bats and their environment has been reported, such as Rabies virus (RABV) and Histoplasma capsulatum. Thus, the aim of this work was to review the reports of zoonotic pathogens associated with bats in Brazil in the past ten years. We reviewed the main pathogenic microorganisms described and the species of bats most frequently involved in the epidemiological cycles of these zoonotic agents. The obtained data show an upward trend in the detection of zoonotic pathogens in Brazilian bats, such as RABV, Bartonella sp., Histoplasma capsulatum and Leishmania spp., with emphasis on the bat species Artibeus lituratus, Carollia perspicillata, Desmodus rotundus and Molossus molossus. These findings highlight the importance of monitoring bat-associated microrganisms to early identify pathogens that may threaten bat populations, including potentially zoonotic microrganisms, emphasizing the importance of the One Health approach to prevent and mitigate the risks of the emergence of zoonotic diseases., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Cationic glycopolymers stand out as gene delivery nanosystems due to their inherent biocompatibility and high binding affinity to the asialoglycoprotein receptor (ASGPR), a target receptor overexpressed in hepatocellular carcinoma (HCC) cells. However, their synthesis procedure remains laborious and complex, with problems of solubilization and the need for protection/deprotection steps. Here, a mini-library of well-defined poly(2-aminoethyl methacrylate hydrochloride- co -poly(2-lactobionamidoethyl methacrylate) (PAMA- co -PLAMA) glycopolymers was synthesized by activators regenerated by electron transfer (ARGET) ATRP to develop an efficient gene delivery nanosystem. The glycoplexes generated had suitable physicochemical properties and showed high ASGPR specificity and high transfection efficiency. Moreover, the HSV-TK/GCV suicide gene therapy strategy, mediated by PAMA 144 - co -PLAMA 19 -based nanocarriers, resulted in high antitumor activity in 2D and 3D culture models of HCC, which was significantly enhanced by the combination with small amounts of docetaxel. Overall, our results demonstrated the potential of primary-amine polymethacrylate-containing-glycopolymers as HCC-targeted suicide gene delivery nanosystems and highlight the importance of combined strategies for HCC treatment.

This study aimed to standardize the use of an ex vivo wound model for the evaluation of compounds with antibiofilm activity. The in vitro susceptibility of Staphylococcus aureus ATCC 29213 and Pseudomonas aeruginosa ATCC 27853 to ciprofloxacin and polyhexamethylene biguanide (PHMB) was evaluated in planktonic and biofilm growth. The effects of ciprofloxacin and PHMB on biofilms grown on porcine skin explants were evaluated by colony-forming unit (CFU) counting and confocal microscopy. Minimum inhibitory concentrations (MICs) against S. aureus and P. aeruginosa were, respectively, 0.5 and 0.25 µg mL-1 for ciprofloxacin, and 0.78 and 6.25 µg mL-1 for PHMB. Minimum biofilm eradication concentrations (MBECs) against S. aureus and P. aeruginosa were, respectively, 2 and 8 µg mL-1 for ciprofloxacin, and 12.5 and >25 µg mL-1 for PHMB. Ciprofloxacin reduced (P < 0.05) log CFU counts of the biofilms grown ex vivo by 3 and 0.96 for S. aureus and P. aeruginosa, respectively, at MBEC, and by 0.58 and 8.12 against S. aureus and P. aeruginosa, respectively, at 2xMBEC. PHMB (100 µg/mL) reduced (P < 0.05) log CFU counts by 0.52 for S. aureus and 0.68 log for P. aeruginosa, leading to an overall decrease (P < 0.05) in biofilm biomass. The proposed methodology to evaluate the susceptibility of biofilms grown ex vivo led to reproducible and reliable results., (© The Author(s) 2023. Published by Oxford University Press on behalf of Applied Microbiology International.)

Trichosporon spp. are emerging opportunistic fungi associated with invasive infections, especially in patients with haematological malignancies. The present study investigated the in vitro inhibition of efflux pumps by promethazine (PMZ) as a strategy to control T. asahii and T. inkin . Planktonic cells were evaluated for antifungal susceptibility to PMZ, as well as inhibition of efflux. The effect of PMZ was also studied in Trichosporon biofilms. PMZ inhibited T. asahii and T. inkin planktonic cells at concentrations ranging from 32 to 256 μg   ml -1 . Subinhibitory concentrations of PMZ inhibited efflux activity in Trichosporon . Biofilms were completely eradicated by PMZ. PMZ potentiated the action of antifungals, affected the morphology, changed the amount of carbohydrates and proteins and reduced the amount of persister cells inside biofilms. The results showed indirect evidences of the occurrence of efflux pumps in Trichosporon and opens a perspective for the use of this target in the control of trichosporonosis.

This study evaluated the effect of the iron chelator deferiprone (DFP) on antimicrobial susceptibility and biofilm formation and maintenance by Burkholderia pseudomallei . Planktonic susceptibility to DFP alone and in combination with antibiotics was evaluated by broth microdilution and biofilm metabolic activity was determined with resazurin. DFP minimum inhibitory concentration (MIC) range was 4-64 µg/mL and in combination reduced the MIC for amoxicillin/clavulanate and meropenem. DFP reduced the biomass of biofilms by 21 and 12% at MIC and MIC/2, respectively. As for mature biofilms, DFP reduced the biomass by 47%, 59%, 52% and 30% at 512, 256, 128 and 64 µg/mL, respectively, but did not affect B. pseudomallei biofilm viability nor increased biofilm susceptibility to amoxicillin/clavulanate, meropenem and doxycycline. DFP inhibits planktonic growth and potentiates the effect of β-lactams against B. pseudomallei in the planktonic state and reduces biofilm formation and the biomass of B. pseudomallei biofilms.

Objectives: To document the work situation, the work ability and the expectation of returning to work among adult patients with systemic autoimmune myopathies (SAMs), and to identify the factors associated with each of these outcomes., Methods: Cross-sectional study. The work situation (performing paid work vs out of work) was ascertained via a structured questionnaire. For those who were working, we applied the Work Ability Index (WAI; scale 7-49); and for those who were out of work, we applied the Return-to-Work Self-Efficacy questionnaire (RTW-SE; scale 11-66)., Results: Of the 75 patients with SAMs included, 33 (44%) were doing paid work and 42 (56%) were out of work. The work situation was independently associated with physical function, assessed by the Health Assessment Questionnaire-Disability Index (HAQ-DI). A 1-point increase in the HAQ-DI (scale 0-3) decreased the chance of doing paid work by 66% (95% CI: 0.16, 0.74; P = 0.007). Patients performing paid work had a mean WAI of 33.5 (6.9). The following variables were associated with a decrease in the WAI score in the regression model: female sex (-5.04), diabetes (-5.94), fibromyalgia (-6.40), fatigue (-4.51) and severe anxiety (-4.59). Among those out of work, the mean RTW-SE was 42.8 (12.4). Cutaneous manifestations and >12 years of education were associated with an average increase of 10.57 and 10.9 points, respectively, in the RTW-SE. A 1-point increase in the HAQ-DI decreased the RTW-SE by 4.69 points., Conclusion: Our findings highlight the poor work participation in a well-characterized sample of working-age patients with SAMs. Strategies to improve work-related outcomes in these patients are urgently needed., (© The Author(s) 2022. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Chimeric antigen receptor T cell (CAR T cell) therapy is a revolutionary approach approved by the FDA and EMA to treat B cell malignancies and multiple myeloma. The production of these T cells has been done through viral vectors, which come with safety concerns, high cost and production challenges, and more recently also through electroporation, which can be extremely cytotoxic. In this context, nanosystems can constitute an alternative to overcome the challenges associated with current methods, resulting in a safe and cost-effective platform. However, the barriers associated with T cells transfection show that the design and engineering of novel approaches in this field are highly imperative. Here, we present an overview from CAR constitution to transfection technologies used in T cells, highlighting the lipid- and polymer-based nanoparticles as a potential delivery platform. Specifically, we provide examples, strengths and weaknesses of nanosystem formulations, and advances in nanoparticle design to improve transfection of T cells. This review will guide the researchers in the design and development of novel nanosystems for next-generation CAR T therapeutics., Competing Interests: Declaration of Competing Interest Henrique Faneca reports a relationship with Lupagen, Inc. that includes: funding grants. Rosemeyre Cordeiro reports a relationship with Gilead Sciences, Lda. that includes: funding grants., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

Copolymers composed of low-molecular-weight polyethylenimine (PEI) and amphiphilic Pluronics® are safe and efficient non-viral vectors for pDNA transfection. A variety of Pluronic® properties provides a base for tailoring transfection efficacy in combination with the unique biological activity of this polymer group. In this study, we describe the preparation of new copolymers based on hydrophilic Pluronic® F68 and PEI (F68PEI). F68PEI polyplexes obtained by doping with free F68 (1:2 and 1:5 w/w) allowed for fine-tuning of physicochemical properties and transfection activity, demonstrating improved in vitro transfection of the human bone osteosarcoma epithelial (U2OS) and oral squamous cell carcinoma (SCC-9) cells when compared to the parent formulation, F68PEI. Although all tested systems condensed pDNA at varying polymer/DNA charge ratios (N/P, 5/1−100/1), the addition of free F68 (1:5 w/w) resulted in the formation of smaller polyplexes (<200 nm). Analysis of polyplex properties by transmission electron microscopy and dynamic light scattering revealed varied polyplex morphology. Transfection potential was also found to be cell-dependent and significantly higher in SCC-9 cells compared to the control bPEI25k cells, as especially evident at higher N/P ratios (>25). The observed selectivity towards transfection of SSC-9 cells might represent a base for further optimization of a cell-specific transfection vehicle.

Persistent apical periodontitis occurs when the endodontic treatment fails to eradicate the intraradicular infection, and is mainly caused by Gram-positive bacteria and yeasts, such as Enterococcus faecalis and Candida albicans , respectively. Phenothiazines have been described as potential antimicrobials against bacteria and fungi. This study aimed to investigate the antimicrobial potential of promethazine (PMZ) and chlorpromazine (CPZ) against E. faecalis and C. albicans dual-species biofilms. The susceptibility of planktonic cells to phenothiazines, chlorhexidine (CHX) and sodium hypochlorite (NaOCl) was initially analyzed by broth microdilution. Interaction between phenothiazines and CHX was examined by chequerboard assay. The effect of NaOCl, PMZ, CPZ, CHX, PMZ + CHX, and CPZ + CHX on biofilms was investigated by susceptibility assays, biochemical and morphological analyses. Results were evaluated through one-way ANOVA and Tukey's multiple comparison post-test. PMZ, alone or in combination with irrigants, was the most efficient phenothiazine, capable of reducing cell counts, biomass, biovolume, carbohydrate and protein contents of dual-species biofilms. Neither PMZ nor CPZ increased the antimicrobial activity of CHX. Further investigations of the properties of phenothiazines should be performed to encourage their use in endodontic clinical practice.

Pyometra is one of the most common diseases in adult female dogs, characterized by a suppurative bacterial infection of the uterus with accumulation of inflammatory exudate and a variety of local and systemic clinical manifestations. This study aimed to identify the bacteria within the uterine content and vaginal canal of bitches with pyometra and evaluate their antimicrobial susceptibility and production of virulence factors. Uterine and vaginal content were collected with sterile swabs from 30 bitches diagnosed with pyometra. Bacteria were identified and assessed for their antimicrobial susceptibility and production of virulence factors, including biofilms, siderophores, proteases and hemolysins, both in planktonic and biofilm forms. A total of 82 bacterial isolates (35 uterus, 47 vagina), belonging to 21 species, were identified, with Escherichia coli as the most prevalent species (32/82, 39%). As for susceptibility, 39/79 (49.4%) isolates were resistant to one or more drugs, with resistance proportion among Gram-positive bacteria (87.5%) higher (p < .05) than that observed for Gram-negative bacteria (32.7%). Four coagulase-negative Staphylococcus species were resistant to methicillin. Regarding virulence, the isolates had low production of biofilms, siderophores, proteases and hemolysins, suggesting that the occurrence of pyometra might be more associated with host-related factors than bacterial virulence., (© 2022 Wiley-VCH GmbH.)

Trichosporon asahii and T. inkin are emergent agents of deep-seated and disseminated infections in immunocompromised patients. The present study aimed to investigate the role of extracellular DNA (eDNA) and the enzyme deoxyribonuclease (DNase) on the structure of T. asahii and T. inkin biofilms, as well as to examine their effect on the susceptibility to antifungals. Biofilms reached maturity at 48 h; eDNA concentration in the supernatant increased over time (6 < 24 h < 48h). Exogenous eDNA increased biomass of Trichosporon biofilms at all stages of development, enhanced their tolerance to antifungals and improved their structural complexity. DNase reduced biomass, biovolume and thickness of Trichosporon biofilms, thereby rendering them more susceptibility to voriconazole. The results suggest the relevance of eDNA in the structure and antifungal susceptibility of Trichosporon biofilms and highlight the potential of DNase as adjuvant in biofilm control.

Paraquat (1,10-dimethyl-4,4-bipyridinium dichloride; PQ) is a free-radical producing herbicide that affects cell membranes and can upset the environmental balance of microorganisms present in soil, such as Cryptococcus spp. This study aimed to evaluate the in vitro activity of PQ against Cryptococcus spp. in planktonic and biofilm forms, as well as the protective effect of antioxidant agents against the antifungal effect of PQ and the kinetics of melanin production in response to PQ. Susceptibility to PQ was evaluated by microdilution. Cryptococcus sp. strains exposed to PQ were grown in media with ascorbic acid (AA) and glutathione (GSH). Melanin production was assessed in the presence of l-3,4-dihydroxyphenylalanine (l-DOPA) + PQ. The minimum inhibitory concentration of PQ against Cryptococcus spp. ranged from 8 to 256 µg/mL. Furthermore, PQ reduced biofilm formation. AA and GSH restored the fungal growth of Cryptococcus spp. exposed to PQ. In addition, l-DOPA + PQ delayed melanin production by 24 and 48 h for C . deuterogattii and C. neoformans sensu lato, respectively, suggesting that PQ induces a fitness trade-off in melanin production. Taken together, our data suggest that the antifungal effect of PQ against Cryptococcus spp. possibly exerts selective pressures interfering with biofilm formation and melanin production by these yeasts.

Objective: To review articles that assessed work-related outcomes such as workability, work productivity, presenteeism, absenteeism, sick leave, return to work, and employment status of Brazilian patients with rheumatoid arthritis, systemic lupus erythematosus, systemic sclerosis, Sjögren's syndrome, and systemic autoimmune myopathies., Methods: This study was conducted in Medline databases (PubMed), SciELO, and Lilacs through a combination of descriptors of interest. Studies published until December 2020 were considered in the search strategy., Results: Eight out of 90 articles met the eligibility criteria and were included in this review. The studies are highly heterogeneous. Most of them are cross-sectional, and all of them address rheumatoid arthritis or systemic lupus erythematosus. A common denominator among these studies is the high proportion of patients outside the labor market., Conclusions: In general, the studies show unfavorable labor outcomes and impaired participation in the Brazilian workforce among the samples of patients assessed. There is a need to better understand several topics about Brazilian patients with systemic autoimmune diseases and their work context, as well as to conduct studies focusing on rarer diseases and on the themes of return and reintegration to work.

Enterococcus faecalis is the most important agent of persistent apical periodontitis, and recently, Candida albicans has also been implicated in periapical infections. This study aimed to optimize an in vitro E. faecalis and C. albicans dual-species biofilm protocol for endodontic research. Different physicochemical conditions for biofilm formation were tested. Susceptibility assays to antimicrobials, biochemical composition and an ultra-morphological structure analyses were performed. Reproducible dual-species biofilms were established in BHI medium at 35 °C, for 48 h and in a microaerophilic atmosphere. An increase in biomass and chitin content was detected after vancomycin treatment. Structural analysis revealed that the dual-species biofilm was formed by both microorganisms adhered to the substrate. The proposed protocol could be useful for the study of interkingdom relationships and help to find new strategies against periapical infections.

In recent years, it has been shown that a combination of different antitumour strategies involving distinct therapeutic agents, such as chemical compounds and genetic material, could result in an effective therapeutic activity that is much higher than that obtained by conventionally used individual approaches. Therefore, the main goal of this work was to develop a new hybrid nanosystem based on mesoporous silica nanoparticles and polymers to efficiently transport and deliver drug and plasmid DNA into cancer cells. Moreover, its potential to mediate a combinatorial antitumour strategy involving epirubicin and herpes simplex virus thymidine kinase/ganciclovir (HSV-TK/GCV) gene therapy was evaluated. For this purpose, various cationic polymers were assessed, including poly(β-amino ester) homopolymer, gelatine type A, gelatine type B, and poly(ethylene glycol)-b-poly(2-aminoethyl methacrylate hydrochloride) block copolymer. The obtained results show that using different polymers leads to nanosystems with different physicochemical properties and, consequently, different biological activities. The best formulation was obtained for hybrid nanosystems coated with PEG-b-PAMA. They demonstrated the ability to cotransport and codeliver an anticancer drug and plasmid DNA and effectively mediate the combined antitumour strategy in 2D and 3D tumour cell culture models. In summary, we developed a novel silica- and polymer-based nanosystem able to mediate a dual chemotherapeutic and suicide gene therapy strategy with a much higher therapeutic effect than that obtained through the use of individual approaches, showing its potential for cancer treatment., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Objectives: To compare the perception of disease activity (DA) between adult patients with systemic autoimmune myopathies (SAMs) and their physicians, and analyse possible sources of discordance., Methods: This cross-sectional study included 75 patients with SAMs. Patients and physicians rated the global DA on a 0-10 cm visual analogue scale. A discrepancy score was calculated by subtracting physician assessment from patient assessment. Three groups were defined: (I) no discrepancy: difference within -2.0 to +2.0; (II) negative discrepancy (ND): difference <-2.0 (patient underrated DA in relation to physcian); (III) positive discrepancy (PD): difference >+2.0 (patient overrated DA in relation to physician). Logistic regression was used to identify predictors of discordance., Results: Discordance in patient-physician assessment of DA was found in 21 (28%) cases. ND was observed in 3 (4%), PD in 18 (24%), and no discrepancy in 54 (72%) assessments. Due to the small number, ND cases were excluded from the analysis. PD was associated with older age, personal history of depression, past joint involvement, higher MMT-8 and lower extramuscular DA. In the regression model, for each additional year of age, the chance of PD increases, on average, by 9% (OR 1.09; 95%CI 1.01-1.17, p=0.034). Personal history of depression increases the chance of PD by 829% (OR 9.29; 95%CI 1.52-56.89, p=0.016)., Conclusions: Almost 30% of patients had discordance in DA assessment from their physicians. The majority of them overrated their DA. These patients tend to be older and are more likely to have personal history of depression, past joint involvement, and milder disease.

Lipopeptide biosurfactants (LBs) are biological molecules with low toxicity that have aroused growing interest in the pharmaceutical industry. Their chemical structure confers antimicrobial and antibiofilm properties against different species. Despite their potential, few studies have demonstrated their capability against Malassezia spp., commensal yeasts which can cause dermatitis and serious infections. Thus, the aim of this study was to evaluate the antifungal activity of biosurfactants produced by new strains of Bacillus subtilis TIM10 and B. vallismortis TIM68 against M. furfur and their potential for removal and inhibition of yeast biofilms. Biosurfactants were classified as lipopeptides by FTIR, and their composition was characterized by ESI-Q-TOF/MS, showing ions for iturin, fengycin, and surfactin, with a greater abundance of surfactin. Through the broth microdilution method, both biosurfactants inhibited the growth of clinical M. furfur strains. Biosurfactant TIM10 showed greater capacity for growth inhibition, with no statistical difference compared to those obtained by the commercial antifungal fluconazole for M. furfur 153DR5 and 154DR8 strains. At minimal inhibitory concentrations (MIC-2), TIM10 and TIM68 were able to inhibit biofilm formation, especially TIM10, with an inhibition rate of approximately 90%. In addition, both biosurfactants were able to remove pre-formed biofilm. Both biosurfactants showed no toxicity against murine fibroblasts, even at concentrations above MIC-2. Our results show the effectiveness of LBs in controlling the growth and biofilm formation of M. furfur clinical strains and highlight the potential of these agents to compose new formulations for the treatment of these fungi., (© The Author(s) 2021. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology.)

This study aimed to identify Candida spp. from agricultural soils cultivated with azole fungicides and investigate their susceptibility to clinical (fluconazole, itraconazole, voriconazole, and amphotericin B) and agricultural (tetraconazole and tebuconazole) antifungals in planktonic form. Additionally, Candida biofilm-forming ability and biofilm susceptibility to agricultural antifungals and voriconazole were analyzed. Species identification was performed by phenotypic and molecular assays. The susceptibility of planktonic cells was evaluated by the broth microdilution method. The biofilm metabolic activity was evaluated by the XTT reduction assay. The recovered Candida spp. were identified as C. parapsilosis sensu stricto (n = 14), C. albicans (n = 5), C. tropicalis (n = 2), C. fermentati (n = 1), and C. metapsilosis (n = 2). Minimum inhibitory concentration ranges for clinical and agricultural antifungals were ≤ 0.03-4 μg/mL and 1-128 μg/mL, respectively. Two and one C. albicans strains were considered non-wild type for voriconazole and fluconazole, respectively. All strains were biofilm producers. The minimum biofilm inhibitory concentration ranges for tetraconazole and tebuconazole were 128-> 1024 μg/mL, while for voriconazole was 512-> 1024 μg/mL. In summary, this study shows that non-wild type and azole-resilient biofilm-producing Candida species colonize agricultural soils cultivated with azole fungicides., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Aim: This study aimed to assess an ex situ model of biofilm-associated wounds on porcine skin for the study of Staphylococcus aureus and Pseudomonas aeruginosa biofilms in a host-like environment, after 48 to 120 h of incubation., Material and Results: Ex situ and in vitro biofilms were comparatively analysed. Overall, CFU-counts and matrix quantification yielded significantly (P < 0·05) higher results for ex situ than in vitro biofilms. Confocal microscopy revealed greater (P < 0·05) biomass and thickness at 48-72 h and greater (P < 0·05) robustness at 72 h of growth. S. aureus ex situ biofilms produced less (P < 0·05) siderophore and proteases than in vitro biofilms, while P. aeruginosa ex situ biofilms produced more (P < 0·05) siderophores and less proteases than in vitro biofilms., Conclusions: Biofilms grown ex situ present a greater amount of bacterial cells and polymeric matrix than their in vitro counterparts, reaching maturity at 72 h of growth. Moreover the production of virulence factors differs between ex situ and in vitro biofilms., Significance and Impact of the Study: These findings emphasize the importance of using ex situ biofilm models, once they mimic in vivo conditions. The use of these models brings perspectives for the pursuit of therapeutic alternatives, as tests may be performed in a host-like environment., (© 2021 The Society for Applied Microbiology.)

Clostridioides difficile is a Gram-positive, spore-forming, anaerobic bacillus which is the leading cause of health-care-associated infective diarrhea. The rising incidence of antibiotic resistance in pathogens such as C. difficile makes researches on alternative antibacterial products very important, especially those exploring natural products like propolis. Brazilian Red Propolis, found in the Northeast region of Brazil, is composed by products from regional plants that have the antimicrobial properties. This study aimed to evaluate the in vitro activity of Brazilian Red Propolis (BRP) against C. difficile strains in planktonic and biofilm forms. The susceptibility of four strains of C. difficile to BRP was analyzed by broth microdilution method and vancomycin was included as control drug. BRP-exposed C. difficile cells were evaluated by scanning electron microscopy (SEM). Then, the effects of BRP on growing and mature C. difficile biofilms were also evaluated. BRP minimum inhibitory concentration was 625 μg/mL against all tested strains, while vancomycin MIC range was 0.5-2 μg/mL. SEM showed the loss of homogeneity in bacterial cell wall and cell fragmentation, after BRP-exposure. BRP, at MIC, reduced (P < 0.05) the biomass, matrix proteins and matrix carbohydrates of growing biofilms, and, at 8xMIC, reduced (P < 0.05) the biomass and matrix proteins of mature biofilms. The present study demonstrated that BRP inhibits planktonic growth, damages cell wall, decreases biofilm growth and harms mature biofilms of C. difficile., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier Ltd. All rights reserved.)