Replication of tobacco mosaic virus on endoplasmic reticulum and role of the cytoskeleton and virus movement protein in intracellular distribution of viral RNA.

Little is known about the mechanisms of intracellular targeting of viral nucleic acids within infected cells. We used in situ hybridization to visualize the distribution of tobacco mosaic virus (TMV) viral RNA (vRNA) in infected tobacco protoplasts. Immunostaining of the ER lumenal binding protein (BiP) concurrent with in situ hybridization revealed that vRNA colocalized with the ER, including perinuclear ER. At midstages of infection, vRNA accumulated in large irregular bodies associated with cytoplasmic filaments while at late stages, vRNA was dispersed throughout the cytoplasm and was associated with hair-like protrusions from the plasma membrane containing ER. TMV movement protein (MP) and replicase colocalized with vRNA, suggesting that viral replication and translation occur in the same subcellular sites. Immunostaining with tubulin provided evidence of colocalization of vRNA with microtubules, while disruption of the cytoskeleton with pharmacological agents produced severe changes in vRNA localization. Mutants of TMV lacking functional MP accumulated vRNA, but the distribution of vRNA was different from that observed in wild-type infection. MP was not required for association of vRNA with perinuclear ER, but was required for the formation of the large irregular bodies and association of vRNA with the hair-like protrusions.

Light-dependent sequestration of TIMELESS by CRYPTOCHROME.

Most organisms have circadian clocks consisting of negative feedback loops of gene regulation that facilitate adaptation to cycles of light and darkness. In this study, CRYPTOCHROME (CRY), a protein involved in circadian photoperception in Drosophila, is shown to block the function of PERIOD/TIMELESS (PER/TIM) heterodimeric complexes in a light-dependent fashion. TIM degradation does not occur under these conditions; thus, TIM degradation is uncoupled from abrogation of its function by light. CRY and TIM are part of the same complex and directly interact in yeast in a light-dependent fashion. PER/TIM and CRY influence the subcellular distribution of these protein complexes, which reside primarily in the nucleus after the perception of a light signal. Thus, CRY acts as a circadian photoreceptor by directly interacting with core components of the circadian clock.

Reciprocal regulation between TOC1 and LHY/CCA1 within the Arabidopsis circadian clock.

The interactive regulation between clock genes is central for oscillator function. Here, we show interactions between the Arabidopsis clock genes LATE ELONGATED HYPOCOTYL (LHY), CIRCADIAN CLOCK ASSOCIATED 1 (CCA1), and TIMING OF CAB EXPRESSION 1 (TOC1). The MYB transcription factors LHY and CCA1 negatively regulate TOC1 expression. We show that both proteins bind to a region in the TOC1 promoter that is critical for its clock regulation. Conversely, TOC1 appears to participate in the positive regulation of LHY and CCA1 expression. Our results indicate that these interactions form a loop critical for clock function in Arabidopsis.

Cloning of the Arabidopsis clock gene TOC1, an autoregulatory response regulator homolog.

The toc1 mutation causes shortened circadian rhythms in light-grown Arabidopsis plants. Here, we report the same toc1 effect in the absence of light input to the clock. We also show that TOC1 controls photoperiodic flowering response through clock function. The TOC1 gene was isolated and found to encode a nuclear protein containing an atypical response regulator receiver domain and two motifs that suggest a role in transcriptional regulation: a basic motif conserved within the CONSTANS family of transcription factors and an acidic domain. TOC1 is itself circadianly regulated and participates in a feedback loop to control its own expression.

[Acquired hemophilia A revealing lung cancer]. / Une hémophilie A acquise ayant révélé un cancer du poumon.

Acquired hemophilia A (AHA) is a rare disease, defined by the production of anti-factor VIII antibodies causing disordered hemostasis. It is idiopathic in 50% of cases, but sometimes associated with solid tumors. We report a case where AHA led to the diagnosis of lung cancer. CASE REPORT: An 82-year-old man with spontaneous hematomas on his trunk and extremities, and isolated prolongation of activated partial thromboplastin time was admitted to the emergency room. A severely reduced factor VIII level and a high factor VIII inhibitor title confirmed the diagnosis of AHA. Thoracic computed tomography scan found a suspect lung nodule and biopsy was consistent with a primary lung adenocarcinoma. The patient received recombinant factor VIII, immunosuppressive therapies, and finally lung stereotactic radiotherapy. Thirty months after diagnosis, the patient is in complete remission both from AHA and from his lung cancer. CONCLUSIONS: Acquired hemophilia A is a rare but potentially severe disease, which may be idiopathic or linked to a solid tumor. The severity of AHA depends on both the volume of hemorrhage and the presence of associated diseases.

¿Afecta la hidradenitis supurativa a la calidad de vida del paciente? Una revision de la literatura / Does hidradenitis suppurativa affect the patient's quality of life? A review of the literature

La hidradenitis supurativa es una enfermedad autoinmune crónica debida a la obstrucción de un folículo piloso, que da lugar a la formación de nódulos. Se desconoce la etiología concreta. Algunos factores desencadenantes son los antecedentes familiares, la obesidad, el hábito tabáquico, la ropa ajustada y los factores hormonales e inmunológicos, entre otros. La sintomatología típica es dolor, supuración, prurito y mal olor, y puede llegar a generar cicatrices, fístulas y tunelizaciones. Las localizaciones de las lesiones típicas son en axilas, zona inguinal, perianal y zona mamaria. Cursa con brotes y no es contagiosa. Dependiendo del estadio existen diferentes tratamientos para esta. El diagnóstico se basa en una entrevista sobre los antecedentes familiares, la localización de las lesiones, las manifestaciones clínicas y la alta recurrencia de la enfermedad. Afecta a un 1% de la población mundial, con mayor incidencia a las mujeres. Objetivos: Investigar si la patología afecta a la calidad de vida de los pacientes y averiguar las repercusiones en esta. Metodología: Se realizó mediante una revisión sistemática de los últimos 5 años, tanto de la hidradenitis supurativa como de la pregunta a estudio en las siguientes bases de datos: Google Académico, PubMed, CINAHL, Web of Science, Scopus y Cochrane. Resultados: Los pacientes refieren tener una peor calidad de vida debido a síntomas de la hidradenitis supurativa, como el dolor, prurito y mal olor. Estos afectan en su vida diaria, vida laboral y relaciones sociales, entre otros. Conclusiones: Tras la búsqueda, se afirma la pregunta de estudio. La calidad de vida del paciente con hidradenitis supurativa está disminuida y deteriorada debido a los síntomas que la cursan. Además, conlleva una serie de comorbilidades tanto psicológicas como personales (AU)

Hidradenitis suppurativa is a chronic autoimmune disease due to the obstruction of a hair follicle leading to the formation of nodules. The specific etiology is unknown. Some triggers are family history, obesity, smoking, tight clothing, hormonal and immunological factors, among others. The typical symptomatology is pain, suppuration, itching, bad smell and can generate scars, fistulas and tunnels. Typical lesion locations are in the armpits, inguinal, perianal, and mammary areas. It has outbreaks and is not contagious. Depending on the stage there are different treatments for it. The diagnosis is based on an interview on the family history, location of the lesions, clinical manifestations and high recurrence of the disease. It affects 1% of the world population, with a higher incidence in women. Objectives Investigate whether the pathology affects the quality of life of patients and find out the repercussions on it. Methods: It was carried out through a systematic review of the last five years of both hidradenitis suppurativa and the question under study in the following databases: Google Scholar, PubMed, CINAHL, Web of Science, Scopus and Cochrane Results: Patients reported having a poorer quality of life due to symptoms of hidradenitis suppurativa such as pain, itching and a bad smell. These affect their daily life, work life, social relationships, among others Conclusions: After the search, the study question is affirmed. The quality of life of the patient with hidradenitis suppurativa is diminished and deteriorated due to the symptoms that occur. In addition, it carries a series of psychological and personal comorbidities (AU)

Impact of a stewardship program on bacteraemia in adult inpatients.

OBJECTIVE: Bloodstream infections (BSIs) are associated with considerable morbidity and mortality among inpatients. The aim of this study was to evaluate the impact of a stewardship program on clinical and antimicrobial therapy-related outcomes in patients with bacteraemia. METHODS: Single-centre, before-and-after quasi-experimental study in adult inpatients. Over 1 January 2013 to 31 June 2013 all patients aged 18 years or older with a bacteraemia (interven-tion group, N=200) were compared to a historical cohort (1 Janu-ary 2012 to 31 December 2012) (control group, N=200). RESULTS: Following blood culture results and adjusting for potential confounders, the stewardship program was associated with more changes to antibiotic regimens (adjusted odds ratio [ORa]: 4.6, 95% CI 2.9, 7.4), more adjustments to antimicrobial therapy (ORa: 2.4, 95% CI 1.5, 3.8), and better source control in the first five days (ORa 1.6, 95% CI: 1.0, 2.7). In the subgroup that initially received inappropriate empiric treatment (n=138), the intervention was associated with more antibiotic changes (OR: 3.9, 95% CI: 1.8, 8.5) and a better choice of definitive antimicrobial therapy (OR 2.3 95% CI: 1.2, 4.6). There were also more antibiotic changes in the subgroups with both Gram-negative (OR: 2.8, 95% CI: 1.6, 4.9; n=217) and Gram-positive (OR: 4.6, 95% CI: 1.8, 9.9; n=135) bacteraemia among those receiving the intervention, while the Gram-positive subgroup also received more appropriate definitive antimicrobial therapy (OR: 3.9, 95% CI: 1.8, 8.8). CONCLUSIONS: The stewardship program improved treatment of patients with bacteraemia and appropriateness of therapy.

Double-balloon enteroscopy: a descriptive study of 50 explorations.

AIM: To evaluate the utility of double-balloon enteroscopy for small-bowel disease. DESIGN: A prospective study of 50 consecutive enteroscopies performed from December 2004 to July 2005 to analyze diagnoses and treatments. PATIENTS: 44 patients (33 had undergone a previous capsule endoscopy) with indications for obscure digestive hemorrhage, angiodysplasia, Peutz-Jeghers syndrome, ulcer, suspected Crohn's disease, tumors, and refractory celiac disease. RESULTS: We carried out enteroscopy studies in 44 patients by the oral route and, in 6 additional patients, by both the oral and anal routes. We reached the ileon with the oral route in all cases but one (jejunal stenosis), and in 4 cases out of 7 with the anal route, with an average duration of 73 minutes. We found angiodysplasia in 19 cases, as well as NSAID-related enteropathy, Crohn's disease, diverticulosis, and Waldenström's disease. We performed biopsies in 31% of cases with diagnoses of adenocarcinoma, lymphangiectasia secondary to tumor in celiac disease, and Whipple's disease. We treated 19 patients with angiodysplasia (1 to 20 synchronous lesions) with argon, and 4 patients with polyps using polipectomy (sporadic polyps or Peutz-Jeghers syndrome). A retained capsule in one patient with stenosis was removed. CONCLUSIONS: Double-balloon enteroscopy is a useful and effective technique in the diagnosis and treatment of small intestine diseases, thus complementing capsule endoscopy. More studies are needed to analyze its impact on the management of this condition.

A shortened, 2-hour rifampin test: a useful tool in Gilbert's syndrome.

INTRODUCTION: Diagnosis of Gilbert's disease often involves unnecessary testing and patient anxiety. Rifampin test can support the diagnosis; it has been described in short series and lacks standardization in dose, collection times, result presentation and interpretation. Our objective was to compare the response to oral rifampin in a series of patients with Gilbert's disease, 2 and 4 h after drug administration. PATIENTS AND METHODS: Eighty-nine patients with Gilbert's disease (elevated total bilirubin with no hepatopathy or hemolysis) were recruited. After a basal blood collection, 900 mg rifampin were administered per os and new samples were drawn 2 and 4 h later. Total and esterified bilirubin were measured in every sample. Haptoglobin concentration was also analyzed. RESULTS: When expressed as relative increase with respect to basal values, variations observed 2 h after rifampin intake were all above 15%. A significant correlation (r = 0.902; p = 0.000) was found between relative increases 2 and 4 h after drug administration. No significant variations were found in haptoglobin concentrations. CONCLUSION: Rifampin test is useful in diagnosing Gilbert's disease, but variations in total bilirubin concentrations (basal and post-rifampin) make that no absolute cut-off value can be used. Correlation between 2- and 4-h relative increases suggests that a shortened version could simplify the test.

Norfloxacin decreases bacterial adherence of quinolone-resistant strains of Escherichia coli isolated from patients with cirrhosis.

BACKGROUND: Long-term administration of norfloxacin is recommended for secondary prophylaxis of spontaneous bacterial peritonitis in cirrhosis, but it may be associated with the development of quinolone-resistant bacteria in stools. However, these bacteria rarely cause infections. AIM: To assess bacterial adherence of either quinolone-sensitive or -resistant Escherichia coli obtained from stools of cirrhotic patients, as one of the main virulence factors, and its variations when sub-minimum inhibitory concentration of norfloxacin were added to the medium. METHODS: E. coli strains were co-cultured with oral epithelial cells obtained from patients in presence/absence of norfloxacin. Bacterial adherence was measured as percentage of cells exhibiting positive adherence and the number of bacteria attached to epithelial cells. RESULTS: 37 sensitive and 22 resistant E. coli strains were studied. Bacterial adherence was similar in both series (78% vs. 81%, P = N.S.), and these percentages were similarly and significantly reduced when subminimum inhibitory concentration of norfloxacin was added to the culture medium (P < 0.001). CONCLUSIONS: Bacterial adherence of E. coli obtained from patients with cirrhosis is unrelated to the sensitivity/resistance to quinolones, and is similarly reduced in both cases when subminimum inhibitory concentration of norfloxacin is added to the medium.

Implementation and assessment of a fast-track programme to improve communication between primary and specialized care in patients with suspected cancer: how to shorten time between initial symptoms of cancer, diagnosis and initiation of treatment.

PURPOSE: This study aims to asses a cancer fast-track programme (CFP) to shorten the time since a patient with suspicion of cancer is referred by the primary care (PC) physician to the specialized medical team. METHODS: Guidelines for main suspected tumours were designed to help PC physicians to detect and rapidly refer cases to the CFP oncology coordinator, who sent them to the appropriate department to accelerate diagnosis, staging and therapy. All patients analysed in this report were referred from June 2009 to July 2012. RESULTS: A total of 897 suspected cancer cases were submitted and finally 705 were studied. In 205 (29 %) a cancer diagnosis was confirmed within 23 days (median). Therapy was initiated within 46 days after referral (median). Early diagnoses with a potential curative approach were made in 166 (82 %). CONCLUSIONS: This CFP decreased the waiting time for cancer diagnosis, by improving communication between PC physician and specialized care teams. Most patients included in this program could get therapy with curative intent.

Evaluation of Helicobacter pylori diagnostic methods in patients with liver cirrhosis.

BACKGROUND: Helicobacter pylori-associated peptic ulcer is a frequent complication in cirrhotic patients and its morbidity rate is high. In spite of this, diagnostic methods for H. pylori infection have not been fully evaluated in these patients. AIM: To evaluate H. pylori diagnostic methods in patients with liver cirrhosis. METHODS: One hundred and one cirrhotic patients were included in the study. Three antral and two corpus biopsies were obtained for rapid urease test of the antral mucosa, and Giemsa stain and immunohistochemistry were performed for both the corpus and antrum. Serology, 13C-urea breath test and faecal H. pylori antigen determination were also carried out. RESULTS: Sixty-two patients were positive and 35 were negative for H. pylori infection; four were indeterminate. The sensitivity and specificity were 90.4% and 100%, respectively, for antral histology, 100% and 100% for gastric body histology, 90.4% and 100% for antral immunohistochemistry, 96.2% and 96.7% for body immunochemistry, 85.7% and 97% for rapid urease test, 83.6% and 55.9% for serology, 96.4% and 97.1% for 13C-urea breath test and 75.4% and 94.1% for faecal antigen. CONCLUSION: The most reliable tests for H. pylori infection in cirrhotic patients were the 13C-urea breath test and gastric body histology.

Echovirus 4 and type 1 diabetes mellitus.

AIMS/HYPOTHESIS: To determine the association between exposure to enteroviruses and Type 1 diabetes. METHODS: We measured neutralizing antibodies to the following enteroviruses: Coxsackievirus CA9, CB1, CB2, CB3, CB4, CB5, CB6, and Echovirus E4, E6, E9, E11 in the sera of (1) Type 1 diabetic patients at diagnosis (n = 33), (2) healthy offspring of parents with Type 1 diabetes without islet cell antibodies (ICA) (n = 43) and (3) normal controls (n = 57). All subjects were less than 20 years old. We performed the neutralization test determining the cytopathogenic effect on Vero cells. HLA DR serotyping was also performed in Group 2. RESULTS: Type 1 diabetic patients showed a higher frequency (21.2%, p < 0.01) of neutralizing antibodies to E4 in relation to controls (1.8%), although there were no differences comparing with offspring of Type 1 diabetic patients (20.9%). Healthy offspring carrying Type 1 diabetes HLA DR susceptibility genes were also exposed to E4 (15.0%). High frequencies of neutralizing antibodies to most enteroviruses were found in the control group. CONCLUSION: This study shows the association between Type 1 diabetes and the presence of neutralizing antibodies to Echovirus 4, suggesting the possible participation of this virus as an environmental trigger of this autoimmune disease. Interestingly, our population displays high frequencies of exposure to enterovirus (including CB4) although the incidence for Type 1 diabetes is low (2.9 per 100,000 inhabitants).

Recurring free-floating thrombus in the left atrium in a patient with mitral prosthesis.

A case of recurrent left atrial free-floating thrombus in a patient with mitral prosthesis and multiple emboli is described. Transthoracic echocardiography failed to show the first ball thrombus, whereas transesophageal echocardiography proved highly useful both in diagnosis and in indicating the left atrial appendage as the source of the thrombi.

Non-isotopic tissue-printing hybridization: a new technique to study long-distance plant virus movement.

A non-isotopic tissue-print hybridization technique was developed to study long-distance plant virus movement. By using digoxigenin-labeled RNA probes the distribution pattern of the viral RNA was observed in leaf, stem and petiole tissues. In leaf tissue viral RNA was confined preferentially to symptoms and veins, and in stem and petiole sections, the hybridization signal was observed in vascular tissue. Both chemiluminescent and colorigenic detection methods were used. The colorigenic method, though less sensitive, is advantageous in that it gives some anatomical information on the signal distribution. This non-isotopic tissue-print hybridization technique can provide considerable information about the spatial and temporal virus expression with regard to its symptoms.

Chemiluminescent and colorigenic detection of cherry leaf roll virus with digoxigenin-labeled RNA probes.

Digoxigenin-labeled RNA probes were used to detect cherry leaf roll virus in infected plants. A dot-blot hybridization immunoenzymatic assay in both crude sap extracts and partially purified tissue with a colorigenic and chemiluminescent detection was developed. The use of the new AMPPD substrate was found to be effective in clarified sap extracts in conditions were the colorigenic detection method failed. Both detection assays were effective when using unfractionated nucleic acid preparations, the chemiluminescent being five times more sensitive than the colorigenic. The chemiluminescent hybridization assay makes it possible to detect the virus at the picogram level. The non-radioactive dot-blot hybridization techniques described here turned out to be very suitable for plant virus diagnosis. The sensitivity of this method and those obtained by ELISA or radioactive dot-blot described previously is compared.

Viral isolation from cases of epidemic neuropathy in Cuba.

OBJECTIVE: To investigate the possibility of a viral agent in the central nervous system of patients with epidemic neuropathy. DESIGN: Virus isolation attempts, in cell cultures and suckling mice, from cerebrospinal fluid (CSF) of neuropathy patients and controls undergoing lumbar puncture for unrelated reasons. Serologic studies in patients, contacts, and controls. SETTING: An epidemic of optic and peripheral neuropathy affected more than 50,000 people in Cuba in 1991 through 1993. Illness was associated with dietary limitations and increased physical demands accompanying the shortages of food and fuel experienced in Cuba since 1989. Most patients responded to parenteral vitamin therapy, and the epidemic began to subside when oral vitamin supplementation was begun for the entire Cuban population. RESULTS: Coxsackievirus A9 (five isolates) and a similar, less cytopathic virus (100 isolates) were recovered from 105 (84%) of 125 CSF specimens from neuropathy patients. The strains with light cytopathic effect were antigenically related to Coxsackieviruses A9 and B4 by cross-neutralization and immunoblotting assays. Virus persisted in CSF of some patients for 1 to 12 months. Cerebrospinal fluid from patients and both types of virus from cell culture produced illness, including complete posterior flaccid paralysis, in newborn mice, and virus was reisolated from the mice. Mouse tissues and sural nerve biopsy specimens from patients were stained by immunoperoxidase and colloidal gold techniques using hyperimmune rabbit antisera against the virus with light cytopathic effect. CONCLUSIONS: Coxsackievirus A9 or an antigenically related agent with a light cytopathic effect was present in CSF of 84% of 125 patients with epidemic neuropathy. The role of these agents, probably in combination with nutritional factors, in the pathophysiology of the disease requires further investigation.

Health-risk perception in the inner city community of Centro Habana, Cuba.

Perceptions of health risks were surveyed in the inner city of Centro Habana, Cuba. A questionnaire developed by community leaders and experts was administered to 348 residents to determine the level of perceived risk for each of 41 risk items. Ecologic-level data on morbidity, mortality, and environmental indicators were also gathered. Using factor analysis to reduce the dimensionality of the data, five factor groupings accounted for 60% of the variance, as follows: social environment (40.8%); infectious agents and other health-risk factors of immediate concern (6.1%); lifestyle risks (4. 9%); environmental sanitation (4.1%); and living conditions (3.3%). A relationship between the perception of risk and the ecologic data was found, with inconsistencies largely attributable to factors known to influence risk perception. The greatest concern identified throughout the municipality was housing conditions, highest in the neighborhood that had already begun to address this problem. The analysis was useful in planning targeted health promotion campaigns and prioritizing further interventions. Repeat evaluation of risk perception will be conducted following the completion of interventions.

Platelet dysfunction in cardiopulmonary bypass: an experimental comparative study between a centrifugal and a new pulsatile pump.

The aim of this investigation was to study the effect of a new pulsatile pump for extracorporeal circulation (ECC) on platelet count and platelet function with respect to a Biomedicus centrifugal pump. Thirteen pigs, 8 in the pulsatile group (PG) and 5 in the centrifugal group (CG), underwent a partial extracorporeal circulation lasting 3 h. The animals were sacrificed 3 h post-ECC. The platelet study was both quantitative (platelet count) and qualitative (platelet function analysis) by assessing the closure time (CT) with a PFA-100 system. The decrease in platelet number from basal to 3 h post ECC was only significant in CG (p = 0.009). The platelet function was impaired in both groups, but the value of CT with col/ADP increased significantly only in CG (p < 0.001). The increase of CT with col/EPI was greater in CG (p = 0.07) than in PG (p = 0.2). The results indicated that the new pulsatile pump preserves platelets quantitatively and qualitatively well compared to a Biomedicus pump.

Diaper dermatitis. Value of vitamin A topically applied.

The role of vitamin A in preventing diaper dermatitis was evaluated in a double-blind, randomized, prospective study in which 114 newborns were enrolled over an 11-month period. Patients in Group A (58 infants) were treated with a cream that contained 1,000 IU/g of vitamin A, whereas patients in Group B (56 infants) were treated with a cream that had the same composition, but it did not contain vitamin A. Each participant returned once every 15 days for six follow-up visits.