Development of a national proficiency test for SARS-CoV-2 detection by PCR in Colombia.

Background: Proficiency testing (PT) is a tool for ensuring the validity of results of testing laboratories and is essential when laboratories are working with assays authorised for emergency use or implementing novel techniques for detecting emerging pathogens. Methods: In collaboration with the National Health Institute of Colombia and with international support, we developed a qualitative PT for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by reverse transcription polymerase chain reaction (RT-PCR). A proficiency test item (PTI) based on reference material (research grade) produced by the National Institute of Standards and Technologies (NIST) was prepared and characterised using three positive samples with varying concentrations of SARS-CoV-2 ribonucleic acid (RNA) and two negative (control) samples. Tests were distributed to 121 laboratories across the national network of public health laboratories in Colombia. Results: Positive samples had varying concentrations of SARS-CoV-2 RNA and were quantified by digital PCR (RT-ddPCR) assays for the E gene of SARS-CoV-2. We tested the ability of laboratories to detect low and high levels of viral RNA using samples with SARS-CoV-2 RNA concentrations of 1417 ± 216, 146 ± 28, and 14 ± 10 copies /uL (expanded uncertainty, k = 2, 95% confidence level) We also performed a semiquantitative analysis of instrumental responses (Ct values) reported by participating laboratories and homogeneity, stability, and characterisation studies of the produced materials to determine the adequacy of these materials and methods for use in the qualitative PT scheme. The PT evaluated reports for individual target genes from each laboratory; 98.3% of laboratories had satisfactory performance and the remaining 1.7% of laboratories had unsatisfactory performance for the detection of at least one of the reported genes. Conclusions: This PT scheme identified the potential metrological weaknesses of laboratories in the detection of SARS-CoV-2 by RT-PCR and may facilitate improvements in the quality of measurements from the perspective of public health surveillance.

Grace Under Pressure: Resiliency of Quality Monitoring of Stroke Care During the Covid-19 Pandemic in Mexico City.

Stroke is one of the leading causes of death and disability among adults worldwide. The World Health Organization (WHO) officially declared a COVID-19 pandemic on March 11, 2020. The first case in Mexico was confirmed in February 2020, subsequently becoming one of the countries most affected by the pandemic. In 2020, The National Institute of Neurology of Mexico started a Quality assurance program for stroke care, consisting of registering, monitoring and feedback of stroke quality measures through the RES-Q platform. We aim to describe changes in the demand for stroke healthcare assistance at the National Institute of Neurology and Neurosurgery during the pandemic and the behavior of stroke quality metrics during the prepandemic and the pandemic periods. For this study, we analyzed data for acute stroke patients registered in the RES-Q platform, in the prepandemic (November 2019 to February 2020) and pandemic (March-December 2020) periods in two groups, one prior to the pandemic. During the pandemic, there was an increase in the total number of assessed acute stroke patients at our hospital, from 474 to 574. The average time from the onset of symptoms to hospital arrival (Onset to Door Time-OTD) for all stroke patients (thrombolyzed and non-thrombolyzed) increased from 9 h (542 min) to 10.3 h (618.3 min) in the pandemic group. A total of 135 acute stroke patients were enrolled in this registry. We found the following results: Patients in both groups were studied with non-contrast computed tomography (NNCT), computed tomography angiography (CTA), magnetic resonance angiography (MRA), digital subtraction angiography (DSA) or more frequently in the pandemic period (early carotid imaging, Holter monitoring) as needed. Treatment for secondary prevention (antihypertensives, antiplatelets, statins) did not differ. Frequency of performing and documenting the performance of NIHSS scale at arrival and early dysphagia test improved. There was an increase in alteplase use from 21 to 42% (p = 0.03). There was a decrease in door to needle time (46 vs. 39 min p = 0.30). After the implementation of a stroke care protocol and quality monitoring system, acute stroke treatment in our institution has gradually improved, a process that was not thwarted during the COVID-19 pandemic.

Stroke Among SARS-CoV-2 Vaccine Recipients in Mexico: A Nationwide Descriptive Study.

BACKGROUND AND OBJECTIVES: Information on stroke among severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines remains scarce. We report stroke incidence as an adverse event following immunization (AEFI) among recipients of 79,399,446 doses of 6 different SARS-CoV-2 vaccines (BNT162b2, ChAdOx1 nCov-19, Gam-COVID-Vac, CoronaVac, Ad5-nCoV, and Ad26.COV2-S) between December 24, 2020, and August 31, 2021, in Mexico. METHODS: This retrospective descriptive study analyzed stroke incidence per million doses among hospitalized adult patients (≥18 years) during an 8-month interval. According to the World Health Organization, AEFIs were defined as clinical events occurring within 30 days after immunization and categorized as either nonserious or serious, depending on severity, treatment, and hospital admission requirements. Acute ischemic stroke (AIS), intracerebral hemorrhage (ICH), subarachnoid hemorrhage (SAH), and cerebral venous thrombosis (CVT) cases were collected through a passive epidemiologic surveillance system in which local health providers report potential AEFI to the Mexican General Board of Epidemiology. Data were captured with standardized case report formats by an ad hoc committee appointed by the Mexican Ministry of Health to evaluate potential neurologic AEFI against SARS-COV-2. RESULTS: We included 56 patients (31 female patients [55.5%]) for an overall incidence of 0.71 cases per 1,000,000 administered doses (95% CI 0.54-0.92). Median age was 65 years (interquartile range [IQR] 55-76 years); median time from vaccination to stroke (of any subtype) was 2 days (IQR 1-5 days). In 27 (48.2%) patients, the event was diagnosed within the first 24 hours after immunization. The most frequent subtype was AIS in 43 patients (75%; 0.54 per 1,000,000 doses, 95% CI 0.40-0.73), followed by ICH in 9 (16.1%; 0.11 per 1,000,000 doses, 95% CI 0.06-0.22) and SAH and CVT, each with 2 cases (3.6%; 0.03 per 1,000,000 doses, 95% CI 0.01-0.09). Overall, the most common risk factors were hypertension in 33 (58.9%) patients and diabetes in 22 (39.3%). Median hospital length of stay was 6 days (IQR 4-13 days). At discharge, functional outcome was good (modified Rankin Scale score 0-2) in 41.1% of patients; in-hospital mortality rate was 21.4%. DISCUSSION: Stroke is an exceedingly rare AEFI against SARS-CoV-2. Preexisting stroke risk factors were identified in most patients. Further research is needed to evaluate causal associations between SARS-COV-2 vaccines and stroke.

Microvessel-on-Chip Fabrication for the In Vitro Modeling of Nanomedicine Transport.

Tumor-on-chip devices are becoming ideal platforms to recreate in vitro the particular physiological microenvironment of interest for onco-nanomedicine testing and development. This work presents a strategy to produce a round artificial microvessel on-a-chip device for the study of physiologically relevant nanomedicine transport dynamics. The microchannels have a diameter in the range of the tumor capillaries and a semicircular geometry. This geometry is obtained through an intermediate thermal nanoimprint step using a master mold with square-shaped channel structures produced by standard silicon micromachining or by stereolithography three-dimensional (3D) printing. The working microfluidic chip devices are made by casting polydimethylsiloxane on the imprinted intermediate mold. Artificial blood microvessels are created by seeding human endothelial cells into the round-shaped channels acting as the scaffold. The microchip is connected by 3D-printed reservoirs to a pressure controller, allowing for a fine fluidic control. Under physiological flow conditions, the dynamic interaction of nanoparticles (NPs) with the artificial endothelium was assessed by high-magnification fluorescence microscopy. Overtime, internalization of NPs and clustering was observed and the accumulation rate into the endothelial cells could be characterized in real time.

Actin reorganization at the centrosomal area and the immune synapse regulates polarized secretory traffic of multivesicular bodies in T lymphocytes.

T-cell receptor stimulation induces the convergence of multivesicular bodies towards the microtubule-organizing centre (MTOC) and the polarization of the MTOC to the immune synapse (IS). These events lead to exosome secretion at the IS. We describe here that upon IS formation centrosomal area F-actin decreased concomitantly with MTOC polarization to the IS. PKCδ-interfered T cell clones showed a sustained level of centrosomal area F-actin associated with defective MTOC polarization. We analysed the contribution of two actin cytoskeleton-regulatory proteins, FMNL1 and paxillin, to the regulation of cortical and centrosomal F-actin networks. FMNL1 ß phosphorylation and F-actin reorganization at the IS were inhibited in PKCδ-interfered clones. F-actin depletion at the central region of the IS, a requirement for MTOC polarization, was associated with FMNL1 ß phosphorylation at its C-terminal, autoregulatory region. Interfering all FMNL1 isoforms prevented MTOC polarization; nonetheless, FMNL1 ß re-expression restored MTOC polarization in a centrosomal area F-actin reorganization-independent manner. Moreover, PKCδ-interfered clones exhibited decreased paxillin phosphorylation at the MTOC, which suggests an alternative actin cytoskeleton regulatory pathway. Our results infer that PKCδ regulates MTOC polarization and secretory traffic leading to exosome secretion in a coordinated manner by means of two distinct pathways, one involving FMNL1 ß regulation and controlling F-actin reorganization at the IS, and the other, comprising paxillin phosphorylation potentially controlling centrosomal area F-actin reorganization. ABBREVIATIONS: Ab, antibody; AICD, activation-induced cell death; AIP, average intensity projection; APC, antigen-presenting cell; BCR, B-cell receptor for antigen; C, centre of mass; cent2, centrin 2; cIS, central region of the immune synapse; CMAC, CellTracker™ Blue (7-amino-4-chloromethylcoumarin); cSMAC, central supramolecular activation cluster; CTL, cytotoxic T lymphocytes; DAG, diacylglycerol; DGKα, diacylglycerol kinase α; Dia1, Diaphanous-1; dSMAC, distal supramolecular activation cluster; ECL, enhanced chemiluminescence; ESCRT, endosomal sorting complex required for traffic; F-actin, filamentous actin; Fact-low cIS, F-actin-low region at the centre of the immune synapse; FasL, Fas ligand; FMNL1, formin-like 1; fps, frames per second; GFP, green fluorescent protein; HBSS, Hank's balanced salt solution; HRP, horseradish peroxidase; ILV, intraluminal vesicles; IS, immune synapse; MFI, mean fluorescence intensity; MHC, major histocompatibility complex; MIP, maximal intensity projection; MVB, multivesicular bodies; MTOC, microtubule-organizing centre; NS, not significant; PBL, peripheral blood lymphocytes; PKC, protein kinase C; PKCδ, protein kinase C δ isoform; PLC, phospholipase C; PMA, phorbol myristate acetate; Pol. Index, polarization index; pSMAC, peripheral supramolecular activation cluster; PSF, point spread function; ROI, region of interest; SD, standard deviation; shRNA, short hairpin RNA; SEE, Staphylococcus enterotoxin E; SMAC, supramolecular activation cluster; TCR, T-cell receptor for antigen; T-helper (Th); TRANS, transmittance; WB, Western blot.

Facial injuries and the gender issue: expressions of violence in a metropolitan region of Northeastern Brazil

Abstract The aims of this investigation were to describe the profile of men and women victims of violence and identify factors associated with the severity of facial trauma. A retrospective study was carried out from 762 records of victims attended at the Institute of Legal Medicine and Dentistry located in a metropolitan region of Northeastern Brazil. The dependent variable was type of facial trauma suffered by victims. Independent variables were the sociodemographic characteristics of victims, characteristics of aggressors and circumstances of violence. Descriptive, bivariate (c2 test) and multivariate statistics were made through logistic regression. Level of significance was set at p < 0.05. The mean age of victims was 29.78 years (SD=13.33). Based on the final regression model, male subjects [odds ratio (OR)=2.22, 95% CI=1.08-4.57, p=0.030], assaulted by other male subjects (OR=4.88; 95% CI=1.12-21.26; p=0.035) through instrument (OR=6.67; 95% CI=2.85-15.60; p<0,001) or mixed aggressions (OR=4.34; 95% CI=1.44-13.02; p=0.009) were more likely to exhibit facial bone fractures or dentoalveolar fractures. The findings highlight that men and women present important victimization differentials in relation to interpersonal violence and facial trauma. Victim's gender, aggressor's gender and mechanism of aggression may exert influence on facial trauma patterns.

Resumo Os objetivos desta investigação foram descrever o perfil de homens e mulheres vítimas de violência e identificar fatores associados à gravidade do trauma facial. Foi realizado um estudo retrospectivo de 762 prontuários de vítimas atendidas no Instituto de Medicina Legal e Odontologia de uma região metropolitana do Nordeste do Brasil. A variável dependente foi o tipo de trauma facial sofrido pelas vítimas. Variáveis ​​independentes foram as características sociodemográficas das vítimas, características dos agressores e circunstâncias de violência. Estatísticas descritivas, bivariadas (teste c2) e multivariadas foram feitas por meio de regressão logística. O nível de significância foi fixado em p<0,05. A idade média das vítimas foi de 29,78 anos (DP =13,33). Com base no modelo de regressão final, os indivíduos do sexo masculino [odds ratio (OR)=2,22, IC 95%=1,08-4,57, p=0,030], agredidos por outros sujeitos do sexo masculino (OR=4,88; IC 95%=1,12-21,26; p=0,035) por meio de instrumentos (OR=6,67; IC 95%=2,85-15,60; p<0,001) ou agressões mistas (OR=4,34; IC 95%=1,44-13,02; p=0,009) foram mais propensos a apresentar fraturas de ossos faciais ou fraturas dentoalveolares. Os achados apontam que homens e mulheres apresentam importantes diferenciais de vitimização em relação à violência interpessoal e trauma facial. O gênero da vítima, o gênero do agressor e o mecanismo de agressão podem exercer influência sobre os padrões de trauma facial.

Protein Kinase C δ Regulates the Depletion of Actin at the Immunological Synapse Required for Polarized Exosome Secretion by T Cells.

Multivesicular bodies (MVB) are endocytic compartments that enclose intraluminal vesicles (ILVs) formed by inward budding from the limiting membrane of endosomes. In T lymphocytes, ILVs are secreted as Fas ligand-bearing, pro-apoptotic exosomes following T cell receptor (TCR)-induced fusion of MVB with the plasma membrane at the immune synapse (IS). In this study we show that protein kinase C δ (PKCδ), a novel PKC isotype activated by diacylglycerol (DAG), regulates TCR-controlled MVB polarization toward the IS and exosome secretion. Concomitantly, we demonstrate that PKCδ-interfered T lymphocytes are defective in activation-induced cell death. Using a DAG sensor based on the C1 DAG-binding domain of PKCδ and a GFP-PKCδ chimera, we reveal that T lymphocyte activation enhances DAG levels at the MVB endomembranes which mediates the association of PKCδ to MVB. Spatiotemporal reorganization of F-actin at the IS is inhibited in PKCδ-interfered T lymphocytes. Therefore, we propose PKCδ as a DAG effector that regulates the actin reorganization necessary for MVB traffic and exosome secretion.

Embarazo espontáneo en una paciente con agenesia de cuello uterino / Spontaneous pregnancy in patient with cervical agenesis

Resumen ANTECEDENTES: La incidencia de agenesia cervical con endometrio funcional se desconoce, aunque se calcula menor a 0.1% en la población general. El pronóstico reproductivo de pacientes con malformaciones müllerianas es limitado y requiere múltiples intervenciones quirúrgicas. CASO CLÍNICO: Paciente primigesta de 21 años, con antecedente de agenesia de cuello uterino y dos tercios superiores de la vagina. Acudió a consulta a las 38.1 semanas de embarazo establecido conforme a la fecha de la última menstruación. Refirió haber concebido de forma espontánea y negó complicaciones durante el embarazo. Se programó para finalizar el embarazo mediante cesárea. El periodo trans y posquirúrgico transcurrió sin complicaciones maternas ni fetales. CONCLUSIÓN: El embarazo espontáneo en pacientes con malformaciones müllerianas debe tratarse a tiempo para asegurar que no surjan complicaciones que pongan en riesgo la vida de la madre y su hijo.

Abstract BACKGROUND: The incidence of cervical agenesis with functional endometrium is unknown, but it's estimated to be less than 0.1% in the general population. The reproductive prognosis in Müllerian malformations is limited and, in most cases, requires multiple surgical interventions to be improved. CASE REPORT: A 21-year-old primiparous patient with a history of agenesis of the cervix and two upper thirds of the vagina. Attended a first-time obstetric appointment at 38.1 weeks of gestation. She refers that the pregnancy was conceived spontaneously and denied complications during pregnancy. A cesarean section was scheduled to end the pregnancy, which had no trans or post-operative maternal-fetal morbidity. CONCLUSIONS: Spontaneous pregnancy in patients with congenital agenesis of the cervix should be addressed in time to ensure a favorable obstetric outcome.

Disruption of Type I Interferon Induction by HIV Infection of T Cells.

Our main objective of this study was to determine how Human Immunodeficiency Virus (HIV) avoids induction of the antiviral Type I Interferon (IFN) system. To limit viral infection, the innate immune system produces important antiviral cytokines such as the IFN. IFN set up a critical roadblock to virus infection by limiting further replication of a virus. Usually, IFN production is induced by the recognition of viral nucleic acids by innate immune receptors and subsequent downstream signaling. However, the importance of IFN in the defense against viruses has lead most pathogenic viruses to evolve strategies to inhibit host IFN induction or responses allowing for increased pathogenicity and persistence of the virus. While the adaptive immune responses to HIV infection have been extensively studied, less is known about the balance between induction and inhibition of innate immune defenses, including the antiviral IFN response, by HIV infection. Here we show that HIV infection of T cells does not induce significant IFN production even IFN I Interferon production. To explain this paradox, we screened HIV proteins and found that two HIV encoded proteins, Vpu and Nef, strongly antagonize IFN induction, with expression of these proteins leading to loss of expression of the innate immune viral RNA sensing adaptor protein, IPS-1 (IFN-ß promoter stimulator-1). We hypothesize that with lower levels of IPS-1 present, infected cells are defective in mounting antiviral responses allowing HIV to replicate without the normal antiviral actions of the host IFN response. Using cell lines as well as primary human derived cells, we show that HIV targeting of IPS-1 is key to limiting IFN induction. These findings describe how HIV infection modulates IFN induction providing insight into the mechanisms by which HIV establishes infection and persistence in a host.

Integrin α4ß1 is necessary for CD4+ T cell-mediated protection against genital Chlamydia trachomatis infection.

Chlamydia trachomatis infection is the most common sexually transmitted bacterial infection in the United States and a significant health burden worldwide. Protection from Chlamydia infection in the genital mucosa is dependent on IFN-γ derived from CD4(+) Th1 cells. These CD4(+) T cells must home successfully to the genital tract to exert their effector function and decrease C. trachomatis burden. Although adhesion receptors expressed by CD4(+) T cells in the genital tract have been characterized, the integrin receptor required for Chlamydia-specific CD4(+) T cell-mediated protection has not been explored. In this study, we demonstrate that C. trachomatis infection of the upper genital tract results in recruitment of Chlamydia-specific CD4(+) T cells robustly expressing the integrin α4ß1. Interfering with α4ß1, but not α4ß7, function resulted in defective CD4(+) T cell trafficking to the uterus and high bacterial load. We conclude that integrin α4ß1 is necessary for CD4(+) T cell-mediated protection against C. trachomatis infection in the genital mucosa. By identifying homing molecules required for successful CD4(+) T cell trafficking to C. trachomatis-infected tissues, we will be better equipped to design vaccines that elicit sterilizing, long-lasting immunity without inducing immune pathologies in the upper genital tract.

Assessing the oral condition of visually impaired individuals attending the Paraíba Institute of the Blind / Avaliação da condição bucal e da ocorrência de manifestações orais em deficientes visuais assistidos no Instituto dos Cegos da Paraíba

Purpose: To evaluate oral conditions in visually impaired individuals at the Paraíba Institute of the Blind. Methods: This observational and transversal study analyzed 80 users of the institute. Participants were given a clinical evaluation encompassing DMFT (index of decayed, missing and filled teeth), SOHI (Simplified Oral Hygiene Index) and SPR (Simplified Periodontal Registry) and the occurrence of oral manifestations. Results: The DMFT index showed that the group is large and more expressive in adult age. The Oral Hygiene Index indicated a deficiency in this group. The most frequent periodontal finding was gingivitis, but children exhibited a healthy periodontium. Oral manifestations and a significant number of injuries to the anterior teeth were also observed, including gingival hyperplasia, severe dental crowding, aphthous ulcerations, bottle caries, fistula, fissured tongue, dental erosion and marked gum recession. Conclusion: The studied population seems to show an increase in the DMFT index with age. Poor oral hygiene may be present due to the lack of visualization of the act of brushing. Gingivitis is the most prevalent periodontal condition, indicating the need for programs to encourage the promotion of oral health.

Objetivo: Avaliar a condição bucal e a ocorrência de manifestações orais em deficientes visuais assistidos no Instituto dos Cegos da Paraíba, Brazil, uma vez que estes usualmente associam a cavidade bucal apenas ao seu aspecto funcional. Metodologia: Estudo observacional e transversal. A amostra foi constituída por 80 usuários do referido instituto, os quais foram examinados clinicamente, avaliando-se os índices CPOD, SOHI (Índice de Higiene Oral), SPR bem como a ocorrência de manifestações bucais. Resultados: O índice CPOD mostrou-se elevado e mais expressivo na faixa etária adulta. O SOHI apresentou-se deficiente. A condição periodontal mais frequentemente encontrada foi a gengivite, porém as crianças exibiram um periodonto saudável. Quanto às manifestações orais, além de um número expressivo de traumatismos em dentes anteriores, também foram observados: hiperplasia gengival, apinhamento dentário severo, ulcerações aftosas, cáries de mamadeira, fistula, língua fissurada, desgastes dentários acentuados e recessão gengival. Conclusão: A população em estudo parece exibir um caráter cumulativo associando-se o índice CPOD ao fator idade; higiene oral deficiente, podendo esta ser justificada pela falta de visualização do ato de escovação; e gengivite como condição periodontal mais frequente, indicando a necessidade de programas de incentivo à promoção de saúde bucal.

Niveles de insulina en recién nacidos macrosómicos hijos de madres no diabéticas / Insulin levels in macrosomic newborns delivered by non diabetic mothers

Objetivo. Describir los niveles de insulina sérica en recién nacidos macrosómicos, hijos de madres no diabéticas. Diseño. Prospectivo, descriptivo, serie de casos encontrados en un período de 6 meses. Población. Veinte recién nacidos con un peso arriba del 90§percentilo para la edad gestacional ysexo. Metodología. Se descartó por historia clínica y niveles normales de glicemia que la madre fuera diabética. Se determinó por radioinmunoensayo los niveles de insulina sérica en todos los recién nacidos. Los controles fueron casos pareados de recién nacidos con peso apropiado para la edad gestacional. Resultados. De los 20 casos, 16 (80) fueron del sexo masculino. El peso promedio fue de 4,320 gms al nacer (4100-4900 gms). Los niveles séricos de insulina fueron estadísticamente superiores en el grupo de estudio (13.73 vrs 0.58 uUl/ml),(p<0.05). Conclusión. Los niveles de insulina de recién nacidos macrosómicos hijos de madres no diabéticas son más altos que los recién nacidos adecuados para su edad gestacional