A library-based approach allows systematic and rapid evaluation of seed region length and reveals design rules for synthetic bacterial small RNAs.

All organisms must respond to environmental changes. In bacteria, small RNAs (sRNAs) are an important aspect of the regulation network underlying the adaptation to such changes. sRNAs base-pair with their target mRNAs, allowing rapid modulation of the proteome. This post-transcriptional regulation is usually facilitated by RNA chaperones, such as Hfq. sRNAs have a potential as synthetic regulators that can be modulated by rational design. In this study, we use a library-based approach and oxacillin susceptibility assays to investigate the importance of the seed region length for synthetic sRNAs based on RybB and SgrS scaffolds in Escherichia coli. In the presence of Hfq we show that 12 nucleotides are sufficient for regulation. Furthermore, we observe a scaffold-specific Hfq-dependency and processing by RNase E. Our results provide information for design considerations of synthetic sRNAs in basic and applied research.

Role of vitrectomy for syphilitic uveitis in a tertiary hospital in Spain.

Diagnosis and treatment of ocular syphilis can be challenging due to the wide spectrum of clinical presentations of this sexually transmitted disease. In some cases of syphilitic panuveitis, pars plana vitrectomy (PPV) can be useful in management since it plays an important role in improving fundus examination allowing treatment of possible retinal associated lesions when vitreous inflammation is intense. We present 3 cases of patients with ocular syphilis that underwent a therapeutic PPV, vitreous sample was taken and analyzed in two of them.

An Optimized Genotyping Workflow for Identifying Highly SCRaMbLEd Synthetic Yeasts.

Synthetic Sc2.0 yeast strains contain hundreds to thousands of loxPsym recombination sites that allow restructuring of the Saccharomyces cerevisiae genome by SCRaMbLE. Thus, a highly diverse yeast population can arise from a single genotype. The selection of genetically diverse candidates with rearranged synthetic chromosomes for downstream analysis requires an efficient and straightforward workflow. Here we present loxTags, a set of qPCR primers for genotyping across loxPsym sites to detect not only deletions but also inversions and translocations after SCRaMbLE. To cope with the large number of amplicons, we generated qTagGer, a qPCR genotyping primer prediction tool. Using loxTag-based genotyping and long-read sequencing, we show that light-inducible Cre recombinase L-SCRaMbLE can efficiently generate diverse recombination events when applied to Sc2.0 strains containing a linear or a circular version of synthetic chromosome III.

DuBA.flow─A Low-Cost, Long-Read Amplicon Sequencing Workflow for the Validation of Synthetic DNA Constructs.

Modern biological science, especially synthetic biology, relies heavily on the construction of DNA elements, often in the form of plasmids. Plasmids are used for a variety of applications, including the expression of proteins for subsequent purification, the expression of heterologous pathways for the production of valuable compounds, and the study of biological functions and mechanisms. For all applications, a critical step after the construction of a plasmid is its sequence validation. The traditional method for sequence determination is Sanger sequencing, which is limited to approximately 1000 bp per reaction. Here, we present a highly scalable in-house method for rapid validation of amplified DNA sequences using long-read Nanopore sequencing. We developed two-step amplicon and transposase strategies to provide maximum flexibility for dual barcode sequencing. We also provide an automated analysis pipeline to quickly and reliably analyze sequencing results and provide easy-to-interpret results for each sample. The user-friendly DuBA.flow start-to-finish pipeline is widely applicable. Furthermore, we show that construct validation using DuBA.flow can be performed by barcoded colony PCR amplicon sequencing, thus accelerating research.

The retinal bacillary layer detachment: Clinical features and outcomes in posterior uveitis.

PURPOSE: To describe the clinical characteristics, presentation and response to treatment in posterior uveitis patients with bacillary layer detachment (BLD) seen on optical coherence tomography (OCT). MATERIALS AND METHODS: Retrospective review of patients with posterior uveitis and SD-OCT scans consistent with BLD. Data collected included demographics, uveitic etiology, treatment and duration of follow-up. Outcome measures included macular volume, central subfoveal thickness and visual acuity. RESULTS: Sixteen patients (20 eyes) were included. Twelve were female (75%). The mean age was 43.68 ± 14.7 years. The most frequent etiology of the uveitis was Vogt-Koyanagi-Harada (VKH) disease (n=10), followed by sympathetic ophthalmia (n=2). BLD was bilateral in four patients. Eight patients were treated with intravenous methylprednisolone boluses. Immunosuppressive therapies were required in 8 patients. The mean follow-up was 70 months (range: 2.0-216.0). CONCLUSION: BLD was observed in a series of posterior uveitis cases of various etiologies, showing functional and structural resolution with treatment in most cases.

Fortuitously compatible protein surfaces primed allosteric control in cyanobacterial photoprotection.

Highly specific interactions between proteins are a fundamental prerequisite for life, but how they evolve remains an unsolved problem. In particular, interactions between initially unrelated proteins require that they evolve matching surfaces. It is unclear whether such surface compatibilities can only be built by selection in small incremental steps, or whether they can also emerge fortuitously. Here, we used molecular phylogenetics, ancestral sequence reconstruction and biophysical characterization of resurrected proteins to retrace the evolution of an allosteric interaction between two proteins that act in the cyanobacterial photoprotection system. We show that this interaction between the orange carotenoid protein (OCP) and its unrelated regulator, the fluorescence recovery protein (FRP), evolved when a precursor of FRP was horizontally acquired by cyanobacteria. FRP's precursors could already interact with and regulate OCP even before these proteins first encountered each other in an ancestral cyanobacterium. The OCP-FRP interaction exploits an ancient dimer interface in OCP, which also predates the recruitment of FRP into the photoprotection system. Together, our work shows how evolution can fashion complex regulatory systems easily out of pre-existing components.

Guidance on brolucizumab management recommendations.

PURPOSE: Brolucizumab, a new generation anti-VEGF, has demonstrated efficacy and safety in AMD in the pivotal HAWK and HARRIER trials. Post-marketing, previously undetected adverse events related to intraocular inflammation have been reported. An independent post hoc review of the pivotal trials puts the rate of IOI at 4.6%. The aim of this paper is to propose a set of recommendations for implementing the management of brolucizumab in clinical practice. METHODS: The recommendations made by the authors are based on their clinical experience, critical review of (i) the pivotal trials, the post-hoc analysis of the Safety Review Committee, (ii), and (iii) the published literature. RESULTS: In the pivotal trials, brolucizumab showed sustained functional gains, superior anatomical outcomes with potentially longer intervals between injections and a well-tolerated overall safety profile. Adverse events reported post-marketing include retinal vasculitis and retinal vascular occlusion. Based on the available information, experts recommend (i) ruling out non-recommended patient profiles (prior history of ORI), (ii) screening the patient prior to each injection to rule out active ORI, (iii) monitoring the patient for early warning signs, and (iv) treating immediately should any adverse events develop. CONCLUSIONS: The adverse events reported are rare, but may be associated with severe and irreversible loss of visual acuity. The recommendations made are intended to facilitate the management of brolucizumab in the routine practice of retinologists, to ensure patient safety and, should any adverse events occur, to minimise their impact on vision.

Transit along the vas deferens results in a high percentage of filiform spermatozoa with compacted chromatin in the rooster (Gallus domesticus).

The present work aimed to evaluate the chromatin compaction of rooster spermatozoa along the male reproductive tract, and to study the vas deferens lining cells, potentially involved in sperm maturation. Chromomycin A3 (CMA3) was used to determine the chromatin compaction of spermatozoa from testis (T), proximal (including epididymis, V1), intermediate (V2) and distal (V3) vas deferens, and ejaculate (E). Six Birchen Leonesa roosters were used. E was obtained in vivo by dorso-ventral massage. V1, V2 and V3 sperm were obtained post mortem (six pairs of vasa deferentia), by flushing. T was obtained by washing the testes, cut in halves. The fixed cells were stained with CMA3 and propidium iodide for flow cytometry assessment. Results showed higher (P P P.

Tuberculosis-Related Uveitis in Patients under Anti-TNF-alpha Therapy: A Case Series.

Tumor necrosis factor inhibitors (anti-TNF) have emerged as an effective treatment in noninfectious uveitis (NIU). Anti-TNF may increase the predisposition to infectious disease as tuberculosis (TB). TB-related uveitis in the context of an uveitogenic concurrent systemic immune-mediated disease under anti-TNF treatment remain a diagnostic challenge, deserving special focus on this rare context. Retrospective chart review of patients on anti-TNF drugs for systemic immune-mediated diseases that developed a multicentric microbiologically confirmed active TB with concurrent intraocular involvement.Three patients were recorded. Screening for TB before starting anti-TNF resulted negative in two patients. The other patient had received anti-tuberculous treatment in the past. All showed a microbiologically confirmed extraocular TB after unexpected atypical reactivation of the uveitis shifting to chronic granulomatous pattern.Specialists should be aware of TB reactivation, even with previous negative screening, when ocular uveitis signs and activity do not match with the expected pattern in a patient on anti-TNF drugs.

[Impact of novel coronavirus infection in patients with uveitis associated with an autoimmune disease: result of the COVID-19-GEAS patient survey]. / Impacto de la infección por el nuevo coronavirus en los pacientes con uveítis asociada a una enfermedad autoinmune: resultado de la encuesta COVID-19-GEAS pacientes.

INTRODUCTION: The objetive of these study is to know the characteristics of COVID-19 in patients with uveitis associated with Systemic Autoimmune Disease (SAD) through telematic survey. MATERIAL AND METHODS: Internal Medicine Society and Group of Systemic Autoimmune disease conducted a telematic survey of patients with SAD to learn about the characteristics of COVID-19 in this population. RESULTS: A total of 2,789 patients answered the survey, of which 28 had a diagnosis of uveitis associated with SAE. The majority (82%) were female and caucasian (82%), with a mean age of 48 years. The most frequent SAEs were Behçet's disease followed by sarcoidosis and systemic lupus erythematosus. 46% of the patients were receiving corticosteroid treatment at a mean prednisone dose of 11 mg/day. Regarding infection, 14 (50%) patients reported symptoms compatible with SARS-CoV-2 infection. RT-PCR was performed on the nasopharyngeal smear in two patients and in one of them (4%) it was positive. CONCLUSIONS: Both asymptomatic and symptomatic COVID-19 patients with ASD-associated UNI had received similar immunosuppressive treatment.

Multimodal imaging in acute idiopathic blind spot enlargement syndrome.

Two cases are reported of acute idiopathic blind spot enlargement syndrome (AIBSE) that presented with peripapillary white spots in the fundus. AIBSE belongs to the acute zonal occult outer retinopathy (AZOOR) complex. Conditions of this AZOOR complex may overlap. Typically, in AIBSE, a peripapillary hyperautofluorescence is seen in the autofluorescence. En-face OCT angiography at the ellipsoid level showed hyper-reflective spots around the optic disk in both cases. One case showed a reversible enlargement of the blind spot in visual field. AZOOR complex is an inflammatory disorder that affects the outer retina, and can now be confirmed with structural optical coherence tomography. In the cases presented, there was a reversible severe loss of the outer retina.

Impact of novel coronavirus infection in patients with uveitis associated with an autoimmune disease: Result of the COVID-19-GEAS patient survey.

INTRODUCTION: The objective of these study is to know the characteristics of COVID-19 in patients with uveitis associated with Systemic Autoimmune Disease (SAD) through telematic survey. MATERIAL AND METHODS: Internal Medicine Society and Group of Systemic Autoimmune disease conducted a telematic survey of patients with SAD to learn about the characteristics of COVID-19 in this population. RESULTS: A total of 2,789 patients answered the survey, of which 28 had a diagnosis of uveitis associated with SAE. The majority (82%) were female and caucasian (82%), with a mean age of 48 years. The most frequent SAEs were Behçet's disease followed by sarcoidosis and systemic lupus erythematosus. 46% of the patients were receiving corticosteroid treatment at a mean prednisone dose of 11 mg/day. Regarding infection, 14 (50%) patients reported symptoms compatible with SARS-CoV-2 infection. RT-PCR was performed on the nasopharyngeal smear in two patients and in one of them (4%) it was positive. CONCLUSIONS: Both asymptomatic and symptomatic COVID-19 patients with ASD-associated UNI had received similar immunosuppressive treatment.

Scleritis and sclerokeratitis associated with IgA vasculitis: A case series.

PURPOSE: To describe a case series of scleritis associated with IgA vasculitis (IgAV) at a tertiary referral center. OBSERVATIONS: Three men with scleritis associated with IgAV were identified: one with anterior scleritis alone, one with anterior scleritis and peripheral ulcerative keratitis (sclerokeratitis), and one with anterior and posterior scleritis. Visual acuity was preserved except from the patient who developed posterior scleritis. Ocular pain was the main symptom at presentation. All patients had a previous history of palpable purpura, but only one was aware of his underlying IgAV. Laboratory results revealed microhematuria and proteinuria with normal urinary ß2 microglobulin levels and negative serum ANCAs. Skin or kidney biopsy demonstrated leukocytoclastic vasculitis or glomerulonephritis with dominant IgA immune deposits. CONCLUSIONS AND IMPORTANCE: Although uncommon, IgAV should be included in the differential diagnosis of anterior scleritis alone or associated with peripheral ulcerative keratitis or posterior scleritis, even in systemically asymptomatic patients. Urinalysis should not be underestimated in assessment of scleritis to detect early stages of glomerular disease. Scleritis may be the first manifestation whose study may lead to the diagnosis of IgAV. Multidisciplinary approach is necessary to prevent irreversible organ damage such as renal failure.

Multidisciplinary care and units for uveitis in the internal medicine departments in Spain: Survey of the Systemic Autoimmune Diseases Group.

OBJECTIVE: To identify the multidisciplinary uveitis units in which internal medicine departments participate in collaboration with ophthalmology departments in Spain. MATERIAL AND METHODS: We conducted a multicentre, observational cross-sectional study that collected information using a structured survey sent by email to 1015 partners of the Systemic Autoimmune Diseases Workgroup of the Spanish Society of Internal Medicine (GEAS-SEMI) from the 1st to the 31st of March 2017. RESULTS: We identified a total of 21 support units/consultations for the ophthalmology departments. Seventeen (81%) of the units were specific internal medicine-ophthalmology consultations, and 5 (24%) units had been created in the past 5 years. A median of 460 patients were assessed per unit by the end of the year. CONCLUSIONS: This study shows, for the first time in Spain, the important and close collaboration between ophthalmologists and internists, especially in highly specialized national reference institutions.

Primary Effusion Lymphoma: A Clinicopathological Study of 70 Cases.

Primary effusion lymphoma (PEL) is a rare type of large B-cell lymphoma associated with human herpesvirus 8 (HHV8) infection. Patients with PEL usually present with an effusion, but occasionally with an extracavitary mass. In this study, we reported a cohort of 70 patients with PEL: 67 men and 3 women with a median age of 46 years (range 26-91). Of these, 56 (80%) patients had human immunodeficiency virus (HIV) infection, eight were HIV-negative, and six had unknown HIV status. Nineteen (27%) patients had Kaposi sarcoma. Thirty-five (50%) patients presented with effusion only, 27 (39%) had an extracavitary mass or masses only, and eight (11%) had both effusion and extracavitary disease. The lymphoma cells showed plasmablastic, immunoblastic, or anaplastic morphology. All 70 (100%) cases were positive for HHV8. Compared with effusion-only PEL, patients with extracavitary-only PEL were younger (median age, 42 vs. 52 years, p = 0.001), more likely to be HIV-positive (88.9% vs. 68.6%, p = 0.06) and EBV-positive (76.9% vs. 51.9%, p = 0.06), and less often positive for CD45 (69.2% vs. 96.2%, p = 0.01), EMA (26.7% vs. 100%, p = 0.0005), and CD30 (60% vs. 81.5%, p = 0.09). Of 52 (50%) patients with clinical follow-up, 26 died after a median follow-up time of 40.0 months (range 0-96), and the median overall survival was 42.5 months. The median OS for patients with effusion-only and with extracavitary-only PEL were 30.0 and 37.9 months, respectively (p = 0.34), and patients with extracavitary-only PEL had a lower mortality rate at the time of last follow-up (35% vs. 61.5%, p = 0.07). The median OS for HIV-positive and HIV-negative patients were 42.5 and 6.8 months, respectively (p = 0.57), and they had a similar mortality rate of 50% at last follow-up. In conclusion, patients presenting with effusion-only versus extracavitary-only disease are associated with different clinicopathologic features. PEL is an aggressive lymphoma with a poor prognosis, regardless of extracavitary presentation or HIV status.

Manejo preoperatorio de los adenomas hipofisarios. Lo que un residente de neurocirugía debe conocer / Preoperative management of pituitary adenomas. What a neurosurgery resident should know

Las lesiones selares son una patología con una incidencia de 3,2 a 4 / 100,000 y una prevalencia de 78 a 94 / 100,000. Un 10% son incidentalomas en la población adulta. Se cree que su prevalencia en el orden mundial actualmente va en aumento.En relación a las manifestaciones clínicas, cabe destacar que es una de las pocas enfermedades que pueden manifestarse tanto por signos y síntomas neurológicos (por ejemplo: hemianopsia bitemporal, síndrome de hipertensión endocraneana debido a hidrocefalia, entre otros), como también por síndromes endocrinológicos (por ejemplo: síndrome de Cushing, acromegalia, amenorrea-galactorrea, infertilidad).Todo paciente debe presentar un estudio clínico-radiológico completo, lo que permitirá un correcto diagnóstico y categorización del mismo.El objetivo del presente trabajo es proporcionar al neurocirujano en formación los conceptos claves que servirán de sustento para el manejo preoperatorio de un paciente con adenoma hipofisario.

Sellar lesions are a pathology with an incidence of 3.2 to 4 / 100.000 and a prevalence of 78 to 94 / 100.000. Normally, 10% of them are incidentalomas and adult patients are in the highest risk group. Because it ́s prevalence in the world is currently increasing, it is of extremely importance to study and understand this pathology. In relation to the clinical manifestations, it should be noted that it is one of the few diseases that can manifest through neurological signs and symptoms like bitemporal hemianopsia, endocranial hypertension syndrome due to hydrocephalus, as well as endocrinological syndromes like Cushing's, acromegaly, amenorrhea-galactorrhea and infertility. One of the most important things to notice is that the treatment success in this pathology comes with the correct diagnosis and characterization of it, for what all patients should have a complete clinical-radiological evaluation.In this study, we establish a guide with concepts and key tools to support the medical personal during a pre-surgical preparation of patients with pituitary adenoma.