Novel PCR primers for improved detection of Mycobacterium leprae and diagnosis of leprosy.

AIMS: Diagnosis of leprosy, a chronic infection caused by Mycobacterium leprae, predominantly depends on clinical manifestations and histopathological analysis, hampering rapid and accurate diagnostics. Our aim was to increase accuracy of leprosy diagnosis by improving M. leprae's DNA detection based on polymerase chain reaction (PCR) technique using new specific primers for the RLEP repetitive sequence. METHODS AND RESULTS: The specific target region, RLEP, of M. leprae's genome was selected based on comparative genomics. After confirming the specificity of this region, using blastn analysis, primers were designed and tested for their in silico specificity. To evaluate the specificity and sensitivity of these primers in vitro, 184 blood samples from patients were used in qPCR. The new primer pair LYON1/LYON2 produced 91% positive samples, whereas the current primer pair LP1/LP2 produced 46%. Specificity and DNA detection limit test were carried out to compare the efficiency of the developed primer pair. The LYON1/LYON2 primer showed 100% specificity, whereas LP1/LP2 showed 64%. The DNA detection limit of LYON1/LYON2 was 10 copies of bacterial genomes per millilitre, whereas LP1/LP2 was 1000 copies of bacterial genomes per millilitre. CONCLUSIONS: In conclusion, the developed LYON1/LYON2 primer pair presented to be a specific and sensitive new molecular marker for the diagnosis of leprosy. SIGNIFICANCE AND IMPACT OF THE STUDY: The development of a specific primer pair for the detection of the M. leprae genome through qPCR technique contributes to a fast, sensitive and specific diagnosis, which is essential to prevent spreading and progression of this disease.

Analysis of two meteorite fragments (lunar and martian) using X-Ray microfluorescence and X-Ray computed microtomography techniques.

Meteorites have been arousing the curiosity of mankind since antiquity. However, the interest in these objects goes far beyond mere curiosity in the study of such materials, which has great importance due essentially to the information they can provide. The importance of studying meteorites is associated about the earliest conditions and processes during the formation and earliest history of the solar system. So, in this study, the characterization of two meteorite fragments was performed using X-ray computed microtomography (micro-CT) and X-ray microfluorescence (micro-XRF). These techniques were used for their non-destructive characteristics and the ability to provide information about the structure and composition the meteorites. The micro-CT images showed encrusted structures within both samples. However, while in Lunar meteorites spheroidal structures very similar to small grains internally grouped in clusters were found, in the Martian meteorite a very peculiar structure was identified. Besides that, the micro-CT it was also possible to evaluate the different density materials that compose the samples. The micro-XRF results accounted for the presence of the elements Si, Ca, Ti, Cr, Mn, Fe, Ni and Sr in the Lunar sample, as well as of Si, S, K, Ca, Ti, Cr, Mn, Fe, Cu, Zn, Sr and Y in the Martian sample. The results obtained are effective for the characterization of meteorites, proving thus that it is possible to obtain important information about the chemical composition, as well as about the distribution and the internal structure of these materials, evaluating aspects such as density and porosity.

The Contribution of Inflammation to Autism Spectrum Disorders: Recent Clinical Evidence.

Autism comprises a complex and heterogeneous spectrum of neurodevelopmental disorders, usually termed autism spectrum disorders (ASD). It is more prevalent in males than females, and genetic and environmental factors are believed to account in similar percentages to the development of ASD. In recent years, the contribution of inflammation and inflammatory mediators to disease aetiology and perpetuation has been the object of intense research. In this chapter, inflammatory aspects that contribute to ASD are discussed, including abnormal microglia activation and polarization phenotypes, increased systemic levels of pro-inflammatory mediators, and altered patterns of immune cell response to activation stimuli. Also, inflammation in the context of gut microbiome and the impact of inflammation on gender prevalence of ASD are considered. Finally, treatment impact on inflammatory parameters and the potential for use of anti-inflammatory drugs, alone or in combination with antipsychotics, to manage ASD are examined.

Analysis of metallic archaeological artifacts by x-ray computed microtomography technique.

Metallic archeological artifacts have specific characteristics that differs them to the other classes of materials due to the transformations that occurs to them, mainly electrochemical. This way, a process called corrosion takes place, starting at the surface of the artifacts, changing the metals in oxides, sulphates, carbonates or other compounds that may result in substantial modifications in their morphology or the total mineralization. Possibly, most part of the archeological artifacts was already suffering this natural deterioration process, even before the period of time that they have remained buried. A number of techniques can be used for the analysis of these artifacts; however, most of them require physical sectioning, which is a complex time-consuming process. This work proposes the use of X-ray microtomography (microCT), which is a non-invasive and expeditious analysis technique, in order to identify the structures and texture of metallic archeological artifacts under incrustations. A set of artifacts found in excavations in the city of Rio de Janeiro (Brazil) were investigated. They show evident forms of degradation, and the incrustations did not allow to clearly identifying them. Through the microCT results is was possible to evaluate the structure of the artifacts, as well as to quantify the material loss caused by corrosion. Such results provided significant data about these historic pieces, which is important for a precise diagnostic about their conservation state, being a useful tool for preserving the morphology of the collected artifact.

X-ray microtomography system for small and light samples using a flat panel detector.

A low-cost system able to perform microtomography of samples such as teeth, insects, or other small materials and low atomic numbers is presented. For this, a small flat panel type sensor was used. The process of characterization of the detector is detailed, as well as its main characteristics. The electromechanical control and the software used are also described. The advantages, some limitations, and comparisons with commercial systems are presented along with some three-dimensional volumetric reconstruction of different materials that served as samples during the development of the system.

The bone degenerative processes in senile fishes from Holocene Brazilian shell mounds.

Zooarchaeological collections from shell mounds in Rio de Janeiro (2,470-4,632 cal BP) contain a high prevalence of swollen fish bones belonging to the Atlantic spadefish (Chaetodipterus faber), crevalle jack (Caranx hippos) and fat snook (Centropomus parallelus). Given the lack of knowledge of the bone degenerative process in senile fishes, this study analysed hyperostotic bone in zooarchaeological and modern specimens to obtain high-resolution morphology and microstructure reconstruction. We used microCT as well as X-ray diffraction to characterize the crystallographic changes associated with fish senility. Our results showed that trabecular microstructures in hyperostotic bones were consistent with estimated values of the per cent bone volume-to-total volume ratio (BV/TV) and were greater than 60% in cortical bone. Hyperostotic bones indicated a high radiograph density, and X-ray diffractograms showed a decrease in hydroxyapatite [Ca10 (PO4 )6 (OH)2 ] and calcite (CaCO3 ) neocrystallization. These crystalline and density changes revealed an advanced stage of fish senile and indicate the vulnerability of ageing fish populations.

Genetic diversity of Bromeliaceae species from the Atlantic Forest.

The Bromeliaceae family includes a range of species used for many purposes, including ornamental use and use as food, medicine, feed, and fiber. The state of Espírito Santo, Brazil is a center of diversity for this family in the Atlantic Forest. We evaluated the genetic diversity of five populations of the Bromeliaceae family, including specimens of the genera Aechmea, Billbergia (subfamily Bromelioideae), and Pitcairnia (subfamily Pitcairnioidea), all found in the Atlantic Forest and distributed in the state of Espírito Santo. The number of alleles per locus in populations ranged from two to six and the fixation index (F), estimated for some simple sequence repeats in bromeliad populations, was less than zero in all populations. All markers in the Pitcairnia flammea population were in Hardy-Weinberg equilibrium (P < 0.05). Moreover, significant deviations from Hardy-Weinberg equilibrium were observed at some loci in populations of the five bromeliad species. In most cases, this can be attributed to the presence of inbreeding or the Wahlund effect. The genetic diversity indices of five species showed greater allelic richness in P. flammea (3.55). Therefore, we provide useful information for the characterization of genetic diversity in natural populations of Aechmea ramosa, Aechmea nudicaulis, Billbergia horrid, Billbergia euphemia, and P. flammea in Atlantic Forest remnants in the south of Espírito Santo state.

Treatment of KPC-producing Enterobacteriaceae: suboptimal efficacy of polymyxins.

Treatment of Klebsiella pneumoniae carbapenemase-producing Enterobacteriaceae infections (KPC-EI) remains a challenge. Combined therapy has been proposed as the best choice, but there are no clear data showing which combination therapy is superior. Our aim was to evaluate the effectiveness of antimicrobial regimens for treating KPC-EI. This was a retrospective cohort study of KPC-EI nosocomial infections (based on CDC criteria) between October 2009 and June 2013 at three tertiary Brazilian hospitals. The primary outcomes were the 30-day mortality for all infections and the 30-day mortality for patients with bacteraemia. Risk factors for mortality were evaluated by comparing clinical variables of survivors and nonsurvivors. In this study, 118 patients were included, of whom 78 had bacteraemia. Catheter-related bloodstream infections were the most frequent (43%), followed by urinary tract infections (n = 27, 23%). Monotherapy was used in 57 patients and combined treatment in 61 patients. The most common therapeutic combination was polymyxin plus carbapenem 20 (33%). Multivariate analysis for all infections (n = 118) and for bacteremic infections (n = 78) revealed that renal failure at the end of treatment, use of polymyxin and older age were prognostic factors for mortality. In conclusion, polymyxins showed suboptimal efficacy and combination therapy was not superior to monotherapy.

New microsatellite markers for the neotropical malaria vector Anopheles nuneztovari sensu lato.

Anopheles nuneztovari sensu lato consists of cryptic species and genetic lineages, one of which is an important human malaria vector in the northern part of South America. Population structure and evolutionary genetics studies may help in the definition and delimitation of the species and lineages within this species complex, which is relevant information for organizations involved in malaria control efforts. In this study, 10 new microsatellite markers were isolated from 2 repeat-enriched genomic libraries of A. nuneztovari s.l. and were characterized in 37-48 mosquitoes of this species. All loci were highly polymorphic and encompassed 5-25 alleles per locus. The observed (HO) and expected (HE) heterozygosities ranged from 0.354 to 0.866 and from 0.613 to 0.932, respectively. Six of the 10 new loci showed significant deviations from the Hardy-Weinberg equilibrium, and no linkage disequilibrium was detected. The loci described in this study were more polymorphic than the 18 previously characterized loci and appear to be promising markers for use in investigating the fine-scale population genetic structure and the boundaries of the cryptic species and lineages within the A. nuneztovari complex.

[Assessment of arterial occlusive disease with Duplex sonography. Prospective study]. / Avaliação da doença oclusiva das artérias renais com Triplex Scan. Estudo prospectivo.

Occlusive disease involving the renal arteries represent a major cause of curable hypertension and improvement of renal dysfunction. Angiography is still the major diagnostic procedure. Isotopic methods with Captopril and split-function studies have shown high incidence of false-positive and false-negative results. Colour flow Duplex-Scan renders possible the visualization of the aorta and renal arteries, is noninvasive and combines morphologic and hemodynamic evaluation. To assess its efficacy in the diagnosis of renal artery occlusive disease, a prospective blind study was undertaken in 24 patients, 48 renal arteries, 15 women and 9 men, with mean age of 44 years. Scans of the aorta and renal arteries at the origin, along its extension and at the renal hilum were obtained using and ATL Ultrmark 9 with 3.0 MHZ probe Velocity measurements were obtained at the aorta and along the renal arteries. Criteria for a positive examination were: focal acceleration with a reduction of the flow velocity at the hilum. Absence of flow in a properly visualized renal artery was considered as occlusion. All patients had digital subtraction arteriography. In 1 patient the non-invasive examination was inadequate. In the remain 23 patients the exam was considered adequate. False-positives were 4.8%; false-negatives 8.7%. Sensitivity, specificity and accuracy were respectively 91.7%, and 95.7% and 93.5%. These results suggest that non-invasive assessment of renal artery occlusive disease is feasible and accurate when compared to angiography, thus providing a useful tool for screening hypertensive patients.

Pulmonary findings in patients with chagasic megaesophagus. Study of autopsied cases.

We studied retrospectively 69 cases of chagasic megaesophagus (group 1) and 207 cases of chronic chagasic myocardiopathy (CCM) without megaesophagus (group 2) by autopsy in the Pathology Department of the Federal University of Bahia, Brazil, from 1953 to 1975. It was shown that pulmonary tuberculosis was significantly more frequent in group 1 (21.7 percent) than in group 2 (3.4 percent). The patients of group 1, who had megaesophagus without associated cardiomyopathy (n = 36), had a much higher rate of pulmonary tuberculosis (36.1 percent) than those with associated CCM (6.0 percent), and they also had the most severe forms of the pulmonary disease. Other pulmonary complications of infectious origin occurred more frequently in group 1. The prevalence of pneumonia was 34.7 percent in group 1 and 10 percent in group 2. Six cases were considered to be aspiration pneumonia. Long abscess occurred in two cases in each group. We suggest that megaesophagus should be carefully investigated for pulmonary complications, especially for tuberculosis in populations with a high rate of this infection.