A more reliable PCR for detection of Mycobacterium tuberculosis in clinical samples

Diagnostic techniques based on PCR have two major problems: false-positive reactions due to contamination with DNA fragments from previous PCRs (amplicons) and false-negative reactions caused by inhibitors that interfere with the PCR. We have improved our previously reported PCR based on the amplification of a fragment of the Mycobacterium tuberculosis complex-specific insertion element IS6110 with respect to both problems. False-positive reactions caused by amplicon contamination were prevented by the use of uracil-N-glycosylase and dUTP instead of dTTP. We selected a new set of primers outside the region spanned by the formerly used primers to avoid false-positive reactions caused by dTTP-containing amplicons still present in the laboratory. With this new primer set, 16 copies of the IS6110 insertion element, the equivalent of two bacteria, could be amplified 10(10) times in 40 cycles, resulting in a mean efficiency of 77% per cycle. To detect the presence of inhibitors of the Taq polymerase, which may cause false-negative reactions, part of each sample was spiked with M. tuberculosis DNA. The DNA purification method using guanidinium thiocyanate and diatoms effectively removed most or all inhibitors of the PCR. However, this was not suitable for blood samples, for which we developed a proteinase K treatment followed by phenol-chloroform extraction. This method permitted detection of 20 M. tuberculosis bacteria per ml of whole blood. Various laboratory procedures were introduced to reduce failure or inhibition of PCR and avoid DNA cross contamination. We have tested 218 different clinical specimens obtained from patients suspected of having tuberculosis. The samples included sputum (n=145), tissue biopsy samples (n=25), cerebrospinal fluid (n=15), blood (n=14), pleural fluid (n=9), feces, (n=7), fluid from fistulae (n=2), and pus from a wound (n=1). The results obtained by PCR were consistent with those obtained with culture, which is the "gold standard." We demonstrate that PCR is a useful technique for the rapid diagnosis of tuberculosis at various sites.

Heart failure in young adults

Heart failure (HF) is a major health concern affecting 15 million people in Europe and around 900 000 people in the U.K. HF predominantly affects the elderly, with the mean age of patients with a diagnosis of HF between 70 and 80 years. Most previous HF studies have accordingly focused on older patients. Although HF is less common in younger adults (<65 years), 15% to 20% of patients hospitalised with HF are younger than 60 years of age. Very few studies have described the characteristics of younger adults with HF and its outcome. The aims of this thesis are to describe the clinical characteristics of younger adults with HF, explore the epidemiology of HF in younger adults and determine their short- and long-term outcomes. This was made possible by access multiple databases consisting of large patient cohorts with HF. The first chapter is a systematic literature review of younger adults with HF. Gaps in the current literature were identified and the thesis focused on some of these. The CHARM study allows detail characterisations of younger adults with HF. It recorded characteristics of patients with HF, including symptoms and signs of HF, electrocardiographic changes, chest radiographic findings, and also left ventricular ejection fraction. HF hospitalisations and its precipitating factors were also recorded systematically. Younger adults were more likely to have a third heart sound and hepatomegaly, but less likely to have pulmonary crackles and peripheral oedema. Similarly, radiological findings in younger adults were less likely to show interstitial pulmonary oedema or pleural effusion. Interestingly, younger adults aged <40 years not only have similar HF hospitalisation rate to older patients, however during their presentation with decompensated HF, they were less likely to have clinical pulmonary oedema and radiological signs of HF. Physicians managing younger adults with HF need to be aware of this. Younger adults were also less compliant with medications and lifestyle restriction resulting in hospitalisation with decompensated HF. Fortunately, despite these challenges, mortality rates in younger adults with HF were lower compared to older patients. To further substantiate the findings from the CHARM study, the MAGGIC study, a meta-analysis consists of over 40 000 patients with HF from large observational studies and randomised controlled trials, was examined. In both databases, the commonest aetiology of HF in younger adults was dilated cardiomyopathy. The ejection fraction was the lowest in younger adults. Similar to the CHARM study, mortality rates in younger adults were lower compared to older patients. However, in the MAGGIC study, by stratifying mortality into patients with preserved ejection fraction and with reduced ejection fraction, younger patients with preserved ejection fraction have a much lower mortality rate compared to patients with reduced ejection fraction. Findings from clinical trials are not always reflective of the real life clinical practice. The U.K. Clinical Practice Research Datalink (CPRD), a large and well-validated primary care database with 654 practices contributing information into the database representing approximated 8% of the U.K. population, is a rich dataset offering a unique opportunity to examine the characteristics, treatments, and outcomes of younger adults with HF in the community. In contrast to the CHARM and MAGGIC studies, younger adults aged <40 years were stratified into 20-29 and 30-39 years in the CPRD analysis. This is possible due to the larger number of younger adults with HF. Further stratifying the younger age groups demonstrated heterogeneity among younger adults with HF. In contrast to previous data showing younger adults have lower co-morbidities, the proportions of depression, chronic kidney disease, asthma, and any connective tissue disease were high among patients aged 20-29 years in the analysis from the CPRD. Surprisingly, the treatment rates for angiotensin converting enzyme (ACE) inhibitor, and aldosterone antagonist were the lowest in patients aged 20-29 years. With the exception of patients aged ≥80 years, treatment rate with beta-blocker was also the lowest in patients aged 20-29 years. With over two decades of follow up, long-term mortality rates in younger adults with HF can be determined. The mortality rates continued to decline from 1988 to 2011. Physicians managing younger adults with HF can now use this contemporary data to provide prognostic information to patients and their family. A hospital administrative database is the logical next platform to explore younger adults with HF. The Alberta Ministry of Health database links an outpatient database to a hospitalisation database providing ample data to examine the relationship between outpatient clinic visits and hospital admissions in younger adults with HF. Following a diagnosis of HF in the outpatient setting, younger adults were admitted to the hospital with decompensated HF much sooner than older patients. Younger adults also presented to emergency department more frequently following their first hospitalisation for HF. In conclusion, this thesis presented the characteristics and outcomes of younger adults with HF, and helped to extend our current understanding on this important topic. I hope the data presented here will benefit not only physicians looking after younger adults with HF, but also patients and their family.

Vasculitis, a Rare Presentation of a Post-Streptococcal Syndrome

Post-streptococcal vasculitis is not awell-recognized non-suppurative complication of Group A streptococcal infections. Seventeen-year-old male was admitted with palpable purpura in the limbs, ankle swelling, microscopic hematuria and elevated inflammation markers. After short remission period, an extensive exudative pleural effusion was identified. High anti-streptococcal antibodies titers were found. This report illustrates an atypical and rare presentation of a post-streptococcal syndrome.

TAFRO Syndrome in a Patient of South-American Descent

A 31-year-old Caucasian woman of South-American descent was diagnosed with a variant of multicentric Castleman disease (MCD) that has been reported in Japan as Castleman-Kojima disease. This is a systemic inflammatory disorder known as TAFRO Syndrome which includes thrombocytopenia, polyserositis (ascites/pleural effusion), microcytic anemia, myelofibrosis, fever, renal dysfunction and organomegaly, with immunologic disorder, polyclonal hypergammaglobulinemia, and elevated levels of interleukin-6 (IL-6) and the vascular endothelial growth factor present in serum and/or effusions. Optimal therapies are not well established. The patient was treated with methylprednisolone and rituximab. Following the start of treatment, the patient has been asymptomatic for over 8 months.

Frecuencia y clase de derrame paraneumónico en neumonías adquiridas en la comunidad: hospital Vicente Corral Moscoso 2002-2003

La neumonía adquirida en la Comunidad (NAC) tiene una influencia de 7 a 15 casos por 1000 personas y por año, de los cuales el 36 a 57se acompaña de Derrame Pleural Paraneumónico (DPPN). En el Hospital Vicente Corral Moscoso (HVCM) en el período 1999-2000, la NAC representó el 17de todos los egresos del Departamento de Clínica, correspondiendo un 6.5a DPPN, diagnosticados. Este estudio desciptivo prospectivo realizado en el Departamento de Clínica del HVCM desde septiembre de 2002 a agosto de 2003 muestra que de 1393 pacientes, 127 correspondieron a NAC, y de éstos el 28.3(36 pacientes) fueron diagnosticados de DPPN, las edades estuvieron entre los 20 y 93 años (media 55.4 años), siendo más frecuente en varones. De acuerdo a la clasificación propuesta por Light, los derrames de Clase 1 estuvieron presentes en mayor porcentaje (50.0) y los de clase 7 los menos frecuentes (2.8). El tipo de DPPN determina la conducta terapeútica a seguir es así que el conocimiento de la frecuencia real en el HVCM y su clasificación deberá permitir tomar medidas adecuadas con el fin de mejorar el pronóstico de los pacientes con NAC

A Case of Mesenteric Pseudocyst Causing a Massive Abdominal Swelling

We present the case of a 55-year-old man, with a self-limited febrile condition associated to polyserositis with inconclusive investigation. Bilateral pleural and pericardial effusions resolved. The peritoneal fluid loculated and was compatible with an exudate. The patient remained clinically asymptomatic. Two years later, examination revealed a palpable and painless abdominal mass, which imaging study suggested a cystic lesion. Surgical resection was performed and the histological examination revealed a mesenteric pseudocyst. Mesenteric pseudocysts are rare intra-abdominal cystic masses, mostly benign, without causing specific symptoms. Although imaging tests are useful for their differential diagnosis, the histology is mandatory.

Bilateral Chylothorax Secondary to Retrosternal Goitre: a Case Report and Review of the Literature

Chylothorax is characterized by an accumulation of lymphatic fluid in the pleural cavity due to damage to the thoracic duct. The aetiology can be traumatic or non-traumatic. Goitre is a rare cause of chylothorax with only eight cases previously described in the literature including only one case causing a bilateral chylothorax. This report describes a patient with bilateral chylothorax secondary to substernal goitre, which was successfully treated, and discusses this very rare case in light of the available literature.

Estudo de ß-glucanas extraídas dos fungos Geastrum saccatum e Polyporos dermoporus: características químicas e atividades biológicas

Fungal polysaccharides have received a great deal of attention due to itsbecause of their potential use in a wide rangegreat variety fromof industries. Some studies have demonstrated that polysaccharides extracted offrom basidiomycetes they have presented significant properties as anti-inflammatory, antimicrobial, antioxidant and anti-tumoral properties. In spite of thisDespite these potential properties, these mushrooms have not been insufficiently investigated, and the great number of antibiotics number produced forby these organisms suggests that they canmay be a new source of bioactives composites source. In tThe present work, reports onlated the chemical composition, potential antioxidant, antiinflammatory and citotoxycity of extracted polymers extracted offrom the fruits bodies of the fungiius Geastrum saccatum and Polyporus dermoporus, native mushrooms of the Atlantic forest inof the state of the Rio Grande do Norte, Brazil. The Cchemical analyses had revealed ademonstrated text of total sugar rates of 65% and 49%, and proteins of 7.0% for in extracts of G. saccatum and P. dermoporus extracts, respectively. The analyses ofNMR spectroscopy of RMN had demonstrated that these extracts are composites forof a complex involving β- glucans and- proteins complex. The inhibition of the formation of superoxide radicals formation was of 88.4% in G. saccatum and 83.3% in P. dermoporus, and 75 and 100% for inhibition of hydroxyls radicals inhibition. TopicalThe topic application of extracts the 10, 30 and 50 mg/kg extract in BALBc mice with cutaneous inflammation induced byfor croton oil demonstrated to inhibitedion of ear edema of ear and cells polimorfonuclears cells atin the inflamed siteplace, being this reply more effective in lower concentrations being more effective. The evaluation of the glucans of G. saccatum and P. dermoporus glucans under induced pleurisy for carrageenan-induced pleurisya of showed the antiinflammatory action of these composites., being analyzed tThe frame number in the pleural exudates and thedosage of nitric oxide dosage was also analyzed. The cytotoxic action of these polymers was analyzed throughthrough the mitochondrial function (MTT). The incubation of the glucans with mononuclear cells of the peripheral blood demonstrated that the extracted glucans extracted fromof G. saccatum havepossess a moderate cytotoxic action. These results suggest that these mushrooms possess polymers formed byfor a complex glucana-protein complex, with antiinflammatory and antioxidant actions

Estudo comparativo de carragenanas comerciais Kappa, Iota e Lambda no processo inflamatório em ratos :edema intraplantar e pleurisia

The Iota, Kappa and Lambda commercial carrageenans are rarely pure and normally contain varying amounts of the other types of carrageenans. The exact amount of impurity depends on the seaweed source and extraction procedure. Then, different analysis methods have been applied for determination of the main constituents of carrageenans because these three carrageenans are extensively used in food, cosmetic and pharmaceutical industry. The electrophoresis of these compounds proved that the carrageenans are constituted by sulfated polysaccharides. These compounds were characterized by colorimetric methods and was observed that the Lambda carrageenan shown the greater value (33.38%) of sulfate. These polymers were examined by means of 13C NMR spectroscopy and infrared spectra. The polysaccharides consisted mainly of units alternating of sulfated galactoses and anhydrogalactoses. The aim of the study was also to test the inflammatory action of these different polysaccharides. A suitable model of inflammation is acute sterile inflammation of the rat hind limb induced by carrageenan. Paw edema was induced by injecting carrageenans (κ, ι and λ) in saline into the hind paw of a male Wistar rats (175–200 g). The pathway to acute inflammation by carrageenan (kappa, iota and lambda) were expressed as time-edema dependence and measured by paw edema volume. For this purpose, was used an apparatus (pakymeter), which makes it possible to measure the inflammation (swelling of the rat foot) with sufficient accuracy. The results showed that κ-carrageenan (1%) have an edema of 3.7 mm and the paw edema increase was time and dose dependent; the ι-carrageenan (0.2%) caused an edema of 4 mm and the λ-carrageenan (1%) caused an edema of 3.6 mm. Other model was used in this study based in the inflammation of pleura for comparatives studies. Injection of carrageenans into the pleural cavity of rat induced an acute inflammatory response characterized by fluid accumulation in the pleural cavity, a large number of neutrophils and raised NO production. The levels of NO were measured by Griess reactive. The ι-carrageenan caused the greater inflammation, because it has high concentration of nitrite/nitrate (63.478 nmoles/rat), exudato volume (1.52 ml) and PMNs (4902 x 103 cells). Quantitative evaluation of inflammations of rats is a useful and important parameter for the evaluation of the efficacy of anti-inflammatory drugs

Preparo de amostras citológicas de efusões, lavados (broncoalveolar e peritoneal) e líquor pelo método “plasma-tromboplastina”

Dissertação (mestrado)—Universidade de Brasilia, Faculdade de Ciências da Saúde, Programa de Pós-Graduação em Ciências da Saúde, 2016.

Sequestro pulmonar : trabalho de revisão

Trabalho Final do Curso de Mestrado Integrado em Medicina, Faculdade de Medicina, Universidade de Lisboa, 2014

The Role of Lung Ultrasound in Diagnosis of Respiratory Distress Syndrome in Newborn Infants

Background: Respiratory distress syndrome (RDS) is one of the most common causes of neonatal respiratory failure and mortality. The risk of developing RDS decreases with both increasing gestational age and birth weight. Objectives: The aim of this study was to evaluate the value of lung ultrasound in the diagnosis of respiratory distress syndrome (RDS) in newborn infants. Materials and Methods: From March 2012 to May 2013, 100 newborn infants were divided into two groups: RDS group (50 cases) and control group (50 cases). According to the findings of chest x-ray, there were 10 cases of grade II RDS, 15 grade III cases, and 25 grade IV cases in RDS group. Lung ultrasound was performed at bedside by a single expert. The ultrasound indexes observed in this study included pleural line, A-line, B-line, lung consolidation, air bronchograms, bilateral white lung, interstitial syndrome, lung sliding, lung pulse etc. Results: In all of the infants with RDS, lung ultrasound consistently showed generalized consolidation with air bronchograms, bilateral white lung or alveolar-interstitial syndrome, pleural line abnormalities, A-line disappearance, pleural effusion, lung pulse, etc. The simultaneous demonstration of lung consolidation, pleural line abnormalities and bilateral white lung, or lung consolidation, pleural line abnormalities and A-line disappearance co-exists with a sensitivity and specificity of 100%. Besides, the sensitivity was 80% and specificity 100% of lung pulse for the diagnosis of neonatal RDS. Conclusions: This study indicates that using an ultrasound to diagnose neonatal RDS is accurate and reliable too. A lung ultrasound has many advantages over other techniques. Ultrasound is non-ionizing, low-cost, easy to operate, and can be performed at bedside, making this technique ideal for use in NICU.

Estenosis esofágica. Aplicación de una técnica quirúrgica. Caso clínico

Se presenta el caso clínico de un paciente que presenta estenosis esofágica tras ingesta de caústicos, con disfagia a líquidos por lo que se realizó esofagectomía transhiatal con técnica de tubo gástrico como alternativa para reemplazo esofágico con ascenso mediastinal y yeyunostomía de alimentación, luego del procedimiento presenta fístula a nivel de la anastomosis proximal cervical y derrame pleural que se drena mediante avenamiento pleural bilateral, se maneja de manera conservadora sin desfuncionalización la fistula cervical mediante dren semirígido, con cierre espontaneo y se da de alta al paciente luego de tolerar dieta

Manejo del trauma de tórax y características demográficas, Hospital Vicente Corral Moscoso. Cuenca – Ecuador.

Objetivo: Analizar el manejo del trauma de tórax en el Hospital Vicente Corral Moscoso y describir las características demográficas de la población estudiada. Método: Estudio descriptivo retrospectivo, la muestra fue de 167 pacientes atendidos en el servicio de Emergencia del Hospital Vicente Corral Moscoso, con diagnóstico de trauma de tórax, durante el periodo enero de 2013 a junio de 2015; los datos se obtuvieron de las historias clínicas. Se analizaron las variables de datos demográficos y tratamiento, utilizando frecuencias, porcentajes y chi cuadrado. Se empleó el programa SPSS versión 18, Excel 2010. Resultados: La edad promedio fue de 37 años, con mayor frecuencia en varones con el 84.4%. Los diagnósticos fueron: hemotórax 36.53%, neumotórax 25.75%, hemoneumotórax 27.54%. El 4.8% de los pacientes recibió tratamiento no quirúrgico, el 82.6% avenamiento pleural; al 12.6% se practicó toracotomías al ingreso. Al 19.8% se realizó toracotomía durante la estancia hospitalaria debido a complicaciones como el hemotórax coagulado o residual. La mortalidad fue del 5.4% (9 pacientes), 7 presentaron lesiones extra torácicas. Conclusiones: El trauma de tórax se presentó en alto porcentaje en la tercera década de vida y se resolvió mayormente con avenamiento pleural. Las lesiones extra torácicas incrementan la mortalidad y necesitan un manejo multidisciplinario.

Complications of pneumococcal bacteremia after 13-valent conjugate vaccine withdrawal

In the Region of Madrid, universal immunization with the 13-serotypes pneumococcal conjugate vaccine (PCV13) started in May 2010. In July 2012, public funding ceased. Vaccination coverage decreased from >95% to 82% in 2013 and to 67% in 2014. Our aim was to investigate the impact of PCV13 withdrawal from Madrid Region's universal immunization program on the incidence of complicated pneumococcal bacteremia. We performed a multi-center retrospective cohort study, from 2009 to 2014. Participants were children aged <14 years with Streptococcus pneumoniae bacteremia. Complications were defined as any condition requiring intensive care or surgery. Sequelae were conditions lasting ≥90 days. A total of 168 patients were recruited. One-fourth of both immunized and non-immunized patients had complications. Global complications increased after PCV13 withdrawal. A total of 28% of PCV13 serotypes presented complications. Complications due to PCV13 serotypes did not increase after July 2012. No-PCV13 serotypes increased progressively from 2009 on, and 23% presented complications. A significant risk of complications was found for patients with meningitis, empyema, C-reactive protein >100 mg/L, and serotype 1. A multivariate analysis indicated that complications were associated with meningitis and hospital admission after July 2012. Sequelae were significantly associated with children <2 years of age, meningitis and no-PCV13 serotypes. The incidence of complications due to PCV13 serotypes did not increase two years after PCV13 withdrawal. Nevertheless, all-serotypes complications increased. The likely cause was that no-PCV13 serotypes (associated with meningitis) are on the rise.