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Owing to the rise in multidrug-resistant bacterial diseases and the dwindling supply of newer antibiotics, it is crucial to discover newer compounds or modify current compounds for more effective antimicrobial therapies. According to reports, more than 80% of bacterial infections have been linked to bacterial biofilms. In addition to having antimicrobial properties, the hydrophobic polyphenol curcumin (Cur) also inhibits quorum sensing. The application of curcumin was constrained by its weak aqueous solubility and quick degradation. Over the past years, nanotechnology-based biomaterials with multi-functional characteristics have been engineered with high interest. The present study focused on the development of nano-biomaterials with excellent testifiers for bacterial infection in vitro. In this study, water dispersibility and stability of curcumin were improved through conjugation with gold nanoparticles. The successful synthesis of curcumin-conjugated gold nanoparticles (Cur-AuNPs) was confirmed using X-ray diffraction (XRD), Fourier transform infrared (FTIR) spectroscopy, and UV-vis absorbance spectroscopy. Transmission electron microscopy (TEM) revealed an average particle size of about 10-13 nm. The antibacterial characteristics in terms of the minimum inhibitory concentration (MIC) of Cur-AuNP treatments were found to be lowest than those with AuNPs and Cur treatments. The quantitative analysis revealed the superior antibacterial characteristics of Cur-AuNP-treated bacterial cells compared to the untreated samples. In addition, curcumin-conjugated AuNPs, produced more reactive oxygen species and increased the membrane permeability. Besides, the biocompatibility of Cur-AuNPs was also assessed quantitatively and qualitatively. Statistical analyses revealed the augmented MG-63 cell proliferation in Cur-AuNPs compared to those with Cur and AuNPs treatments. Overall, Cur-AuNPs exhibited enhanced antibacterial, and antibiofilm characteristics and cytocompatibility.
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Curcumina , Nanopartículas Metálicas , Ouro/química , Curcumina/farmacologia , Curcumina/química , Nanopartículas Metálicas/química , Antibacterianos/farmacologia , Antibacterianos/química , ÁguaRESUMO
SARS-CoV-2 is a public pandemic health concern globally. Nasopharyngeal and oropharyngeal swab samples are used for Covid-19 viral detection. Sample collection procedure was tedious and uncomfortable and unsuitable for biochemical and CBC analysis in swab samples. Biochemistry and CBC tests are key determinant in management of Covid-19 patients. We developed a LAMP test to detect viral RNA in blood samples. LAMP is required four specific primers targeting the internal transcribed S-region and loop primers for viral RNA amplification. RNA was extracted from blood samples by TRIzol method. LAMP reaction was performed at 60⯰C for 1 hour and amplicons were visualized in HNB dye. No cross-reactivity was seen with HBV, HCV, and HIV infected sample. Out of 40 blood samples, 33 samples were positive for LAMP and Q-PCR analysis, one sample was positive for LAMP and negative for Q-PCR, two samples were negative for LAMP but positive for Q-PCR, and four blood samples were negative for LAMP and Q-PCR. LAMP method has an accuracy of 92.50%, with sensitivity and specificity of 94.28% and 80%, respectively. Thus, LAMP diagnostic test has proved reliable, fast, inexpensive and can be useful for detection where the limited resources available.â¢LAMP method is a potential tool for detection of SARS-CoV-2.â¢Blood samples are the key determinant for routine diagnostics as well as molecular diagnostics.â¢LAMP assay is an appropriate diagnostics method which offers greater simplicity, low cost, sensitivity, and specificity than other methods in molecular diagnostics.
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Rabies causes approximately 20,000 human deaths in India each year. Nearly all of these occur following dog bites. Large-scale, high-coverage dog rabies vaccination campaigns are the cornerstone of rabies elimination strategies in both human and dog populations, although this is particularly challenging to achieve in India as a large proportion of the dog population are free-roaming and unowned. Further, little is known about free-roaming dog ecology in India which makes defining optimum vaccination strategies difficult. In this study, data collected using a mobile phone application during three annual mass vaccination and neutering (surgical sterilisation of both males and females) campaigns of free-roaming dogs in Ranchi, India (during which a total of 43,847 vaccinations, 26,213 neuter surgeries and 28,172 re-sight observations were made) were interrogated, using two novel approaches to estimate the proportion of neutered dogs that were lost from the city (assumed due to mortality or migration) between campaign years. Analysis revealed high losses of neutered dogs each year, ranging from 25.3% (28.2-22.8) to 55.8% (57.0-54.6). We also estimated that the total population declined by 12.58% (9.89-15.03) over the three-year period. This demonstrates that there is a high turnover of free-roaming dogs and that despite neutering a large number of dogs in an annual sterilisation campaign, the decline in population size was modest over a three-year time period. These findings have significant implications for the planning of rabies vaccination campaigns and population management programmes as well as highlighting the need for further research into the demographics of free-roaming, unowned dogs in India.
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Doenças do Cão , Vacina Antirrábica , Raiva , Animais , Doenças do Cão/epidemiologia , Doenças do Cão/prevenção & controle , Cães , Feminino , Índia/epidemiologia , Masculino , Densidade Demográfica , Dinâmica Populacional , Raiva/epidemiologia , Raiva/prevenção & controle , Raiva/veterinária , Vacinação/veterináriaRESUMO
Background and Purpose: Cerebral aspergillosis is a notorious disease that causes rapid clinical deterioration and carries a poor prognosis. Therefore, it requires timely diagnosis and prompt management. Case Report: This study reports a case of fungal cerebral abscess in a 26years old man following hemodialysis,2 months afterdengue-induced acute kidney disease. Aspergillus fumigatus was recovered from a brain abscess specimen that was subjected to a parietal craniotomy. The patient was successfully treated with oral Voriconazole 400mg BD for 2 days, followed by 200 mg BD for 3months. Conclusion: Hemodialysis patients are at high risk offungal infections due to the frequent use of catheters or the insertion of needles to access the bloodstream. Therefore, a high index of suspicion of fungal infection is required in patients with hemodialysis by the clinician for early diagnosis and treatment.
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We report an experimental study of a Cooper pair splitter based on ballistic graphene multiterminal junctions. In a two transverse junction geometry, namely the superconductor-graphene-superconductor and the normal metal-graphene-normal metal, we observe clear signatures of Cooper pair splitting in the local as well as nonlocal electronic transport measurements. Our experimental data can be very well described by our beam splitter model. These results open up possibilities to design new entangled state detection experiments using ballistic Cooper pair splitters.
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BACKGROUND AND AIMS: Measurement of actual concentration of IgG requires methods like treatment of serum with dithiothreitol (DTT). This study was aimed at comparing of DTT treated ABO titres performed by conventional test tube technique (CTT) and column agglutination technique (CAT) with HA/SPRCA. MATERIALS AND METHODS: This was a prospective, observational study conducted from October 2019 to March 2020. All consecutive A, B and O group donors who gave consent for participation were included. All samples were tested by CTT and CAT before and after DTT treatment (pCTT, pCAT) and with HA/SPRCA. RESULTS: A total of 300 donors were included; 100 each from A, B and O blood group donors. Group O titres were higher than group A/B titres. Group O titres were highest when performed by pCAT, followed by pCTT and lowest by HA/SPRCA. Group A/B titres were highest when performed by HA/SPRCA, followed by pCAT and pCTT for anti-A and highest when performed by pCAT, followed by HA/SPRCA and lowest by pCTT for anti-B. CONCLUSION: Results obtained by pCAT were closer to results obtained by pCTT, whereas those obtained by HA/SPRCA were variable. SPRCA offers the advantage of automation, no inter-observer variation and less time consumption because IgM interference is not observed with SPRCA, thus providing an alternative to pCTT. However, these methods cannot be used interchangeably and to discern the most suitable method, a clinical impact of these results needs to be studied.
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Sistema ABO de Grupos Sanguíneos , Anticorpos , Adesão Celular , Ditiotreitol , Humanos , Estudos ProspectivosRESUMO
OBJECTIVE: To study the clinical profile and factors associated with post-traumatic headache (PTH) in children with mild traumatic brain injury (mTBI) attending an urban public hospital. METHODS: We enrolled 130 consecutive children aged 6-12 years with mTBI (as per the International Classification of Headache Disorders-III criteria), and followed them up for 7 days. Those who developed PTH were further followed up monthly for 3 months. RESULTS: Thirty (23.1%) children developed PTH; 25 (19.2%) children had acute PTH (duration 7 days to 3 months) and the remaining 5 (3.8%) developed persistent PTH (> 3 months). Majority (50%) had bilateral headache and squeezing quality (50%). Forty percent of those with PTH met the criteria for migraine. Obesity (P = 0.84), female gender (P = 0.26), family history of headache (P = 0.93), and prior history of concussion (P = 0.70) were not associated with risk of PTH. Children who developed PTH had higher rate of nausea (RR (95% CI) = 2.42 (1.06, 5.5); P = 0.03) and vomiting (RR (95% CI) = 3.76 (1.64, 8.5); P = 0.001) after mTBI. Headache resolved within 1 month in 63.3% of children. CONCLUSION: PTH was found to be common after mTBI in children. Protocolized follow-up and directed history taking for PTH in all children with mTBI, who are frequently discharge from the emergency department after first aid, will lead to appropriate diagnosis and management of this problem.
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Concussão Encefálica , Cefaleia Pós-Traumática , Concussão Encefálica/complicações , Concussão Encefálica/epidemiologia , Criança , Currículo , Feminino , Cefaleia , Humanos , Cefaleia Pós-Traumática/diagnóstico , Cefaleia Pós-Traumática/epidemiologia , Cefaleia Pós-Traumática/etiologia , Estudos ProspectivosRESUMO
Rice is an important staple food grain consumed by most of the population around the world. With climate and environmental changes, rice has undergone a tremendous stress state which has impacted crop production and productivity. Plant growth hormones are essential component that controls the overall outcome of the growth and development of the plant. Cytokinin is a hormone that plays an important role in plant immunity and defense systems. Trans-zeatin is an active form of cytokinin that can affect plant growth which is mediated by a multi-step two-component phosphorelay system that has different roles in various developmental stages. Systems biology is an approach for pathway analysis to trans-zeatin treated rice that could provide a deep understanding of different molecules associated with them. In this study, we have used a weighted gene co-expression network analysis method to identify the functional modules and hub genes involved in the cytokinin pathway. We have identified nine functional modules comprising of different hub genes which contribute to the cytokinin signaling route. The biological significance of these identified hub genes has been tested by applying well-proven statistical techniques to establish the association with the experimentally validated QTLs and annotated by the DAVID server. The establishment of key genes in different pathways has been confirmed. These results will be useful to design new stress-resistant cultivars which can provide sustainable yield in stress-specific conditions.
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Horsegram (Macrotyloma uniflorum (Lam.) Verdc.) is a drought hardy food and fodder legume of Indo-African continents with diverse germplasm sources demonstrating alternating mechanisms depicting contrasting adaptations to different climatic zones. Tissue specific expression of genes contributes substantially to location specific adaptations. Regulatory networks of such adaptive genes are elucidated for downstream translational research. MicroRNAs are small endogenous regulatory RNAs which alters the gene expression profiles at a particular time and type of tissue. Identification of such small regulatory RNAs in low moisture stress hardy crops can help in cross species transfer and validation confirming stress tolerance ability. This study outlined prediction of conserved miRNAs from transcriptome shotgun assembled sequences and EST sequences of horsegram. We could validate eight out of 15 of the identified miRNAs to demonstrate their role in deficit moisture stress tolerance mechanism of horsegram variety Paiyur1 with their target networks. The putative mumiRs were related to other food legumes indicating the presence of gene regulatory networks. Differential miRNA expression among drought specific tissues indicted the probable energy conservation mechanism. Targets were identified for functional characterization and regulatory network was constructed to find out the probable pathways of post-transcriptional regulation. The functional network revealed mechanism of biotic and abiotic stress tolerance, energy conservation and photoperiod responsiveness.
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Adaptação Fisiológica/genética , Fabaceae/genética , Redes Reguladoras de Genes/genética , MicroRNAs/genética , Aclimatação/genética , Secas , Regulação da Expressão Gênica de Plantas/genética , Estudo de Associação Genômica Ampla/métodos , Proteínas de Plantas/genética , Processamento Pós-Transcricional do RNA/genética , Estresse Fisiológico/genética , Transcriptoma/genéticaRESUMO
Ossifying fibromyxoid tumor (OFMT) is a rare fibro-osseous neoplasm. We present a case highlighting the occurrence of an intracranial OFMT masquerading as meningioma on imaging in a 46-year-old gentleman. Brain imaging revealed an extra-axial calcified lesion along the left cerebellar convexity appearing hypointense on T1- and T2-weighted MRI sequences with no post-contrast enhancement, suggestive of a meningioma. An intraventricular colloid cyst was also noted. The lesion, which was presumed to be meningioma, and the colloid cyst were resected in two subsequent operative settings. Histopathological examination of the calcified lesion confirmed the findings of an OFMT. This report aims to inform the physician about intracranial OFMT mimicking meningioma on imaging. In addition, since multiple brain tumors are not very common, the surgeon should always have a suspicion should there be any heterogeneous and peculiar radiological and histopathological characteristics.
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Chryseobacterium indologenes is a non-glucose fermenting Gram-negative bacteria widely distributed in nature. It has been found to cause a variety of infections like nosocomial pneumonia, bacteremia and wound infections usually in immunosuppressed patients and those with indwelling devices. The organism is resistant to a significant number of the commonly prescribed broad spectrum antibiotics. We report a rare case of meningitis due to C. indologenes in a patient diagnosed with medulloblastoma and hydrocephalus with an external ventricular drain in situ. The patient was successfully treated with trimethoprim/sulfamethoxazole combination and external ventricular drain replacement as shown by subsequent sterile cultures.
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AIM: To assess the value of quantitative spleen and liver volume changes in predicting the survival of patients with primary sclerosing cholangitis (PSC). MATERIALS AND METHODS: This institutional review board-approved single-centre study included 89 PSC patients with baseline and follow-up liver imaging studies and laboratory data between 2000 and 2018. Change in spleen, total and lobar liver volumes, and lobar-to-total liver volume ratio was compared between patients with and without adverse outcome (liver transplantation, transplant waiting list, and death). Receiver operating characteristic (ROC) and Kaplan-Meier analysis were performed to identify the volumetric threshold for prediction of outcome and show how these thresholds predict survival, respectively. A p-value of <0.05 was considered statistically significant. RESULTS: The present cohort included 53 men (60%), with mean age of 42 years at baseline. The only volumetric parameters with significant differences in change between patients with and without adverse outcome were spleen volume (p<0.001) and left-to-total liver volume ratio (L/T; p=0.025). The probability of transplant-free survival at 36 months was 59.1% versus 11.9% for patients with spleen volume change <50 ml versus ≥50 ml, respectively (AUC=0.731); and 61.3% versus 13.8% for patients with L/T change <0.04 versus ≥0.04, respectively (AUC=0.638). The patients with changes below the cut-off in both spleen volume and L/T, had a higher probability of transplant-free survival at 36 months (76.8%), compared to those with change at or below the cut-offs in one or both of these two parameters (36.7%, 15%, respectively; p=0.001). CONCLUSION: Spleen volume change and L/T change might be useful biomarkers for prediction of transplant-free survival in patients with PSC.
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Colangiopancreatografia Retrógrada Endoscópica , Colangiopancreatografia por Ressonância Magnética , Colangite Esclerosante/diagnóstico por imagem , Hepatopatias/diagnóstico por imagem , Esplenopatias/diagnóstico por imagem , Adulto , Colangite Esclerosante/mortalidade , Feminino , Humanos , Hepatopatias/mortalidade , Masculino , Tamanho do Órgão , Valor Preditivo dos Testes , Esplenopatias/mortalidade , Taxa de SobrevidaRESUMO
The occurrence of Escherichia coli O157 and Salmonella spp. in solid bovine manure was investigated through a multi-county survey in California. Solid bovine manure samples (n = 91) were collected from 13 dairy farms located in multiple counties in California between June 2016 and August 2017. To quantify pathogens, DNA was extracted from bacteria in manure samples. Afterwards, the prevalence and levels of E. coli O157 and Salmonella spp. in solid bovine manure were determined by real-time quantitative PCR (qPCR). The prevalence of E. coli O157 and Salmonella spp. in solid bovine manure was 15·4 and 6·6% respectively. Escherichia coli O157 and Salmonella spp. levels in positive samples ranged from 3·1 to 5·3 log CFU per g and from positive (the population was <3 log CFU per g) to 5·2 log CFU per g respectively. Surface samples of manure piles had higher prevalence and levels of E. coli O157 and Salmonella spp. than subsurface samples, while no seasonal effects on pathogen occurrence were observed. Our results indicated that solid bovine manure is a source of E. coli O157 and Salmonella spp. and the application of untreated manure as biological soil amendments may pose potential risks to public health. SIGNIFICANCE AND IMPACT OF THE STUDY: Our findings suggested that the presence of Escherichia coli O157 and Salmonella spp. in solid bovine manure may pose potential risks if untreated manure is applied as biological soil amendments. Considering the large-scale sampling used in this study, the observations provide a holistic assessment in terms of pathogen prevalence in solid bovine manure.
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Infecções por Escherichia coli/epidemiologia , Infecções por Escherichia coli/veterinária , Escherichia coli O157/isolamento & purificação , Esterco/microbiologia , Salmonelose Animal/epidemiologia , Salmonella/isolamento & purificação , Animais , California/epidemiologia , Bovinos , Contagem de Colônia Microbiana , Reação em Cadeia da Polimerase , Prevalência , Solo , Microbiologia do SoloRESUMO
AIM: To analyse the change in size on follow-up of hepatic adenomas (HAs) and adenomatosis, and to investigate the relationship of imaging features with size change. MATERIALS AND METHODS: The study included 44 patients (142 lesions) who underwent magnetic resonance imaging (MRI) or computed tomography (CT) for diagnosis and follow-up of HA. The imaging features and percentage change in maximum tumour dimension were observed over a follow-up duration of up to 139 months. RESULTS: With an average follow-up of 43 months, 37% lesions decreased in size, 58% were stable, 4% increased; one lesion regressed completely. Adenomas were stratified into size groups (<3, 3-5, and ≥5 cm). Size change among the three groups was similar (p>0.05). Percent size change was different for lesions followed for ≤12 months (-7.2%) compared with lesions followed for 13-60 months (-20.5%), and those followed for ≥60 months (-23.5%; p<0.05); there was no difference between lesions followed for 13-60 months and ≥60 months (p=0.523). Baseline size and percent size change was similar between the hepatocyte nuclear factor 1α-inactivated HA (HA-H) and inflammatory HA (HA-I) subtype (p>0.05). CONCLUSION: Most adenomas were either stable or regressed on follow-up. Size change was independent of baseline size. After an initial size decrease within 5 years, no further size reduction was noted on extended follow-up. The percent size change in the HA-H and HA-I subtype was similar.
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Adenoma de Células Hepáticas/diagnóstico por imagem , Neoplasias Hepáticas/diagnóstico por imagem , Neoplasias Primárias Múltiplas/diagnóstico por imagem , Adenoma de Células Hepáticas/patologia , Adulto , Feminino , Seguimentos , Humanos , Fígado/diagnóstico por imagem , Fígado/patologia , Neoplasias Hepáticas/patologia , Imageamento por Ressonância Magnética , Masculino , Pessoa de Meia-Idade , Neoplasias Primárias Múltiplas/patologia , Estudos Retrospectivos , Tomografia Computadorizada por Raios X , Adulto JovemRESUMO
Biofloc technology was evaluated with a view to analyse utilization of nitrogenous waste from the effluent and to improve water quality and growth parameters of Penaeus vannamei in intensive culture system. The experiment was carried out in two different treatment outdoor earthen ponds of 0.12â¯ha, one supplemented with carbon source (molasses, wheat and sugar) for biofloc formation and other was feed based control pond with a stocking density of 60 animals m-2 in duplicate for 120 days. Water, sediment and P. vannamei were sampled at regular intervals from the both set of ponds for evaluating physico-chemical parameters, nitrogen content and growth parameters, respectively. A significant reduction in the concentration of total ammonia nitrogen (TAN) and nitrite (NO2-N) were found in the biofloc pond than that of control pond. A significant low level of nitrogen was recorded in the effluents of biofloc pond in comparison to the control. In biofloc system, a significantly elevated heterotrophic bacterial count along with reduction in total Vibrio count was noticed. A significant improvement in the feed conversion efficiency (FCR) and growth parameters of P. vannamei was noticed in the biofloc pond. Growth of P. vannamei in the biofloc pond showed positive allometric pattern with an increased survival. The microbial biomass grown in biofloc consumes toxic inorganic nitrogen and converts it into useful protein, making it available for the cultured shrimp. This improved FCR and reduced the discharge of nitrogenous waste into adjacent environment, making intensive shrimp farming an eco-friendly enterprise.
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Aquicultura , Penaeidae , Qualidade da Água , Animais , Nitrogênio , Lagoas , ÁguaRESUMO
Dairy manure is regularly applied to crop fields as a solid or liquid to improve the soil nutrient status. However, pathogens may survive during manure storage and enter the environment during application. In this study, three storage practices were evaluated to understand the survival patterns of O157:H7 and spp. in dairy manure using a culture-based approach. To replicate common farm manure storage techniques, solid manure was stacked as piles with periodic turning or as static piles without turning, whereas liquid manure (feces, urine, and water) was stored as a slurry in small tanks to simulate lagoon conditions. The and levels in the manure samples were determined for 29 wk. Results showed that there was an initial reduction in bacteria levels in the first month; however, both and managed to survive in the solid manure piles for the full study period. In slurry samples, was not detected after 14 wk, but survived until the end of the experiment at relatively lower levels than in the solid manure piles. Ambient weather and pile size were identified as the main reasons for bacteria survival during the course of the experiment. The outcome of this study is important in terms of understanding pathogen survival in manure piles and slurries prior to their application to crop fields.
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Escherichia coli O157/fisiologia , Listeria/fisiologia , Esterco/microbiologia , Microbiologia do Solo , Gerenciamento de ResíduosRESUMO
PURPOSE: To evaluate the role of simultaneous superior rectus (SR) recession and anterior transposition of inferior oblique (ATIO) muscle in patients with traumatically lost inferior rectus (IR) muscle. METHODS: Six patients with history of ocular trauma, followed by sudden onset vertical diplopia along with marked hypertropia (HT) and limitation of depression in abduction in the affected eye suggestive of IR disinsertion, were included in this prospective study. The patients were treated by simultaneous SR recession and ATIO muscle in the affected eye by limbal conjunctival approach under local anesthesia. RESULTS: Preoperatively, primary position HT of 40-50 (mean 44.16 ± 4.91) prism diopters (PD) was present in all cases which increased to 65-70 (mean 65.83 ± 5.84) PD in down and in the ipsilateral gaze along with marked limitation of depression in abduction and A pattern. On exploration, the IR could not be traced in four cases. Fibrotic muscle sheath with retracted IR was found 10-12 mm away from the limbus in rest of the two patients. ATIO (6.5 mm from the limbus) with simultaneous recession of ipsilateral SR was done under local anesthesia. At 12 weeks postoperatively, three patients were orthophoric in primary position and vertical alignment with in 4-7 PD in primary position was achieved in rest of the three patients. CONCLUSION: Simultaneous SR recession with ATIO seems to be a good alternative to achieve satisfactory vertical alignment for patients with traumatically lost inferior rectus muscle.
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Diplopia/cirurgia , Músculos Oculomotores/cirurgia , Músculos Oculomotores/transplante , Procedimentos Cirúrgicos Oftalmológicos , Estrabismo/cirurgia , Adulto , Diplopia/etiologia , Traumatismos Oculares/complicações , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Estrabismo/etiologia , Resultado do TratamentoRESUMO
The detection of DMMP (dimethyl methylphosphonate, a simulant of nerve agent sarin) was performed by using p-hexafluoroisopropanol phenyl (HFIPP) functionalized graphene (GR) via hydrogen bond interactions. For this, the HFIPP moiety was covalently functionalized on the surface of GR by a diazo reaction. The HFIPP-GR film-modified QCM electrodes were fabricated and their sensing characteristics towards DMMP were investigated. The proposed sensor showed good response towards sensing DMMP vapor at room temperature. In order to see the effect of HFIPP derivatives on DMMP vapor sensing, a comparative study was also conducted with unfunctionalized graphene. The sensitivity and detection limit of the HFIPP-GR sensor against DMMP vapors were 12.24 Hz ppm-1 and 150 ppb respectively. The HFIPP-GR coated sensors showed good selectivity towards sensing DMMP vapors when compared with common organic vapors.
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PURPOSE: Standardization of Aspergillus polymerase chain reaction (PCR) poses two technical challenges (a) standardization of DNA extraction, (b) optimization of PCR against various medically important Aspergillus species. Many cases of aspergillosis go undiagnosed because of relative insensitivity of conventional diagnostic methods such as microscopy, culture or antigen detection. The present study is an attempt to standardize real-time PCR assay for rapid sensitive and specific detection of Aspergillus DNA in EDTA whole blood. MATERIALS AND METHODS: Three nucleic acid extraction protocols were compared and a two-step real-time PCR assay was developed and validated following the recommendations of the European Aspergillus PCR Initiative in our setup. In the first PCR step (pan-Aspergillus PCR), the target was 28S rDNA gene, whereas in the second step, species specific PCR the targets were beta-tubulin (for Aspergillus fumigatus, Aspergillus flavus, Aspergillus terreus), gene and calmodulin gene (for Aspergillus niger). RESULTS: Species specific identification of four medically important Aspergillus species, namely, A. fumigatus, A. flavus, A. niger and A. terreus were achieved by this PCR. Specificity of the PCR was tested against 34 different DNA source including bacteria, virus, yeast, other Aspergillus sp., other fungal species and for human DNA and had no false-positive reactions. The analytical sensitivity of the PCR was found to be 102 CFU/ml. CONCLUSION: The present protocol of two-step real-time PCR assays for genus- and species-specific identification for commonly isolated species in whole blood for diagnosis of invasive Aspergillus infections offers a rapid, sensitive and specific assay option and requires clinical validation at multiple centers.
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Aspergilose/diagnóstico , Aspergillus/classificação , Aspergillus/isolamento & purificação , Técnicas de Diagnóstico Molecular/normas , Reação em Cadeia da Polimerase em Tempo Real/normas , Aspergilose/microbiologia , Aspergillus/genética , DNA Fúngico/genética , DNA Fúngico/isolamento & purificação , Humanos , Técnicas de Diagnóstico Molecular/métodos , RNA Ribossômico 28S/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Sensibilidade e Especificidade , Fatores de Tempo , Tubulina (Proteína)/genéticaRESUMO
Conducting multicentre operational research is challenging due to issues related to the logistics of travel, training, supervision, monitoring and troubleshooting support. This is even more burdensome in resource-constrained settings and if the research includes patient interviews. In this article, we describe an innovative model that uses open access tools such as Dropbox, TeamViewer and CamScanner for efficient, quality-assured data collection in an ongoing multicentre operational research study involving record review and patient interviews. The tools used for data collection have been shared for adaptation and use by other researchers.
Conduire des recherches opérationnelles multicentriques est un défi, particulièrement dans les contextes de ressources limitées, en tenant compte des questions de logistique de déplacement, de formation, de supervision, de suivi et de soutien à la résolution des problèmes; encore plus si cette recherche implique des entretiens avec des patients. Dans cet article, nous décrivons un modèle innovant qui utilise des outils à accès ouvert comme Dropbox, TeamViewer et CamScanner pour un recueil de données efficace et de qualité assurée dans le cadre d'une recherche opérationnelle continue multicentrique impliquant des revues de dossiers et des entretiens avec des patients. Les outils utilisés pour le recueil de données ont été partagés pour l'adaptation et l'utilisation par d'autres chercheurs.
La realización de investigaciones operativas multicéntricas puede ser problemática, sobre todo en los entornos con restricción de los recursos, habida cuenta de las dificultades en la organización de los desplazamientos, la capacitación, la supervisión, el seguimiento y el apoyo a la resolución de problemas; más aun, cuando la investigación precisa entrevistas a los pacientes. En el presente artículo se describe un modelo innovador que utiliza herramientas de libre acceso como las plataformas Dropbox, TeamViewer y CamScanner, con el fin de lograr una obtención de datos eficiente y de calidad garantizada, en una investigación operativa multicéntrica en curso que comporta el examen de las historias clínicas y entrevistas a los pacientes. Se comunican las herramientas utilizadas en la recogida de datos, con la finalidad de que otros investigadores puedan adaptarlas y las apliquen.