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Biomedical subjects

P Saxena

Publications and source records attributed to P Saxena.

At least 19 recordsLinked to original sources
PubMed2006

Rapid and real-time detection of Chikungunya virus by reverse transcription loop-mediated isothermal amplification assay.

The standardization and validation of a one-step, single-tube, accelerated, quantitative reverse transcription (RT) loop-mediated isothermal amplification (RT-LAMP) assay targeting the E1 gene for the rapid and real-time detection of Chikungunya virus (CHIKV) are reported. A linear relationship between the amount of template and time of positivity value over a range of 2 x 10(8) to 2 x 10(2) copies was obtained. The feasibility of CHIKV RT-LAMP for clinical diagnosis was validated with patient serum samples from an ongoing epidemic in Southern India. Optimal assay conditions with zero background were established for the detection of low levels of CHIKV in acute-phase patient serum samples. The comparative evaluation of the RT-LAMP assay with acute-phase patient serum samples demonstrated exceptionally higher sensitivity by correctly identifying 21 additional positive borderline cases that were missed by conventional RT-PCR (P < 0.0001) with a detection limit of 20 copies. The quantification of virus load in patient serum samples was also determined from the standard curve based on their time of positivity and was found to be in the range of 2 x 10(8) to 2 x 10(1) copies. In addition, the field applicability of the RT-LAMP assay was also demonstrated by standardizing SYBR Green I-based RT-LAMP wherein the amplification was carried out in a water bath at 63 degrees C for 60 min, which was followed by monitoring gene amplification with the naked eye through color changes. These findings demonstrated that the RT-LAMP assay is a valuable tool for rapid, real-time detection as well as quantification of CHIKV in acute-phase serum samples without requiring any sophisticated equipment and has potential usefulness for clinical diagnosis and surveillance of CHIKV in developing countries.

Acute Disease↗
PubMed2006

Development and evaluation of reverse transcription-loop-mediated isothermal amplification assay for rapid and real-time detection of Japanese encephalitis virus.

The standardization and validation of a one-step, single-tube accelerated quantitative reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assay is reported for rapid and real-time detection of Japanese encephalitis virus (JEV). The RT-LAMP assay reported in this study is very simple and rapid; the amplification can be obtained in 30 min under isothermal conditions at 63 degrees C by employing a set of six primers targeting the E gene of JEV. The RT-LAMP assay demonstrated exceptionally higher sensitivity compared to that of RT-PCR, with a detection limit of 0.1 PFU. The specificities of the selected primer sets were established by cross-reactivity studies with other closely related members of the JEV serocomplex as well as by evaluation of healthy human volunteers. The comparative evaluation of the RT-LAMP assay for clinical diagnosis with a limited number of patient cerebrospinal fluid samples revealed 85% concordance with conventional RT-PCR, with a sensitivity and a specificity of 100% and 86%, respectively. The concentration of virus in most of the clinical samples was 10(2) to 10(5) PFU/ml, as determined from the standard curve based on the time of positivity in the samples. In addition, the monitoring of gene amplification can also be visualized with the naked eye by using SYBR green I fluorescent dye. Thus, due to easy operation without a requirement of sophisticated equipment and skilled personnel, the RT-LAMP assay reported here is a valuable tool for the rapid and real-time detection of JEV not only by well-equipped laboratories but also by peripheral diagnostic laboratories with limited financial resources in developing countries.

Antibodies, Viral↗
PubMed2006

Comparison of a dipstick dot-ELISA with commercial assays for anti-dengue virus IgM antibodies.

Dengue virus infections have undergone dramatic expansion in range, affecting several tropical and subtropical regions of the world. Dengue virus causes life-threatening complications characterized by dengue hemorrhagic fever and dengue shock syndrome. No standard validated test systems are available for serological diagnosis of dengue virus infection. This creates problems in the diagnosis and proper management of patients. Evaluation of a Defense Research and Development Establishment (DRDE) dengue virus dipstick dot-ELISA test, developed in house, vis-à-vis commercially available immunodiagnostic kits was carried out for detection of IgM antibodies. The DRDE dengue dipstick dot-ELISA was performed on the basis of the dot-ELISA principle. Commercial tests, namely, the Panbio Dengue IgM Capture ELISA system (Panbio, Sinnamon Park, Australia) and Pathozyme Dengue M (Omega Diagnostics, Alva, UK), were performed according to the manufacturers' protocols. Ninety-one serum samples collected from the states of Kerala and Delhi, India during August and November of 2004 were used in the present study. The overall agreement among all three tests was found to be only 72.16% for IgM antibodies. Correlations between the DRDE dipstick dot-ELISA and the Panbio kit, between the DRDE dipstick dot-ELISA and the Pathozyme Dengue M kit, and between the Panbio kit and the Pathozyme Dengue M kit were found to be 96, 93, and 94%, respectively. Although the Panbio kit is widely used in various laboratories in India, the DRDE dipstick dot-ELISA promises to be a useful kit because of its field applicability and comparable sensitivity.

Animals↗
PubMed2004

Diversity of keratinophilic mycoflora in the soil of Agra (India).

Diversity of keratinophilic mycoflora in the soil of Agra was under observation for 1 year (July 2001-June 2002) and isolation of keratinophilic fungi was followed by the hair-baiting method. The frequency of occurrence of keratinophilic fungi in 284 soil samples collected from various hospitals, cattle yards, poultry farms, crop fields and playgrounds was determined, 204 samples (72%) having been found to be positive. A total of 33 species classified into 11 genera (Acremonium, Aspergillus, Chrysosporium, Emmonsia, Geomyces, Keratinophyton, Microsporum, Myceliophthora, Penicillium, Sporotrichum, Trichophyton) were encountered from the soil samples. Sporotrichum spp. were found to be the most dominant species followed by Trichophyton simii. The parameter of keratinophilic fungi found in the samples studied ranged from 62 to 80% where playgrounds yielded the maximum number of species (80%) while the least dominating soil was hospital soil (62%). Among all the baits used maximum fungi occurred on human hairs (82%) followed by chicken feather (74%), wool (61%) and the least on horns (45%). The spectrum of keratinophilic fungi isolated from different sites differed considerably according to the frequency of use by humans.

Fungi↗
PubMed2004

Assessment of the severity of hand injuries using "hand injury severity score", and its correlation with the functional outcome.

INTRODUCTION: Objective assessment of hand injuries is a complex subject. However, an objective assessment, leading to a score, can help in predicting outcome and can be used as a research tool. Campbell and Kay have devised one such score known as "hand injury severity score" or HISS [J. Hand. Surg. [Br.] 21 (3) (1996) 295]. A study on this score has been carried out in our institute. The idea was to see if the hand injury severity score, correlates with the functional outcome as measured by disability arm shoulder and hand score (DASH), after a period of minimum 2 years. METHOD: All the hand injury patients admitted in the hospital were assessed at the time of admission, and objective information was documented on a hand injury documentation form. The form captured all the data required to calculate "hand injury severity score" . A sample of 70 patients admitted during the first 6 months of 1999 was taken and their scores were calculated. The sample was selected such, that it had a reasonable representation of hand injuries of all severities. After obtaining a due approval from ethics committee, all these 70 patients were sent a DASH questionnaire. A total of 23 patients replied. Spearman's rank correlation test was used to analyse the correlation between the severity of hand injury as assessed using HISS, and functional outcome as measured using DASH. Correlations between the outcome and skeletal component score, outcome and motor component scores, and outcome and Integument component scores, were assessed separately. RESULTS: The study shows a statistically significant association ( r = 0.7182, P = 0.000165) between the severities of injury and the functional outcome. The functional outcome also shows a statistically significant association with the severity of injury to skeletal component (r = 0.5151, P = 0.014083) and motor component (r = 0.6797, P = 0.000507). However, the severity of injury to integument component, as measured by HISS, does not show any association with the outcome (r = 0.3571, P = 0.102736). This study supports the overall structure of the HISS. However an improvement in the integument component scoring is required to improve overall accuracy.

Accidents, Occupational↗
PubMed2003

Diagnostic scanning for suspected scaphoid fractures: an economic evaluation based on cost-minimisation models.

OBJECTIVE: To compare, the costs of diagnosing suspected scaphoid fractures using various protocols with various scanning techniques. DESIGN: Analysis of the cost-minimisation models. The models were prepared after an audit of the existing protocol for suspected scaphoid fractures. SETTING: Departments of Accident and Emergency and Orthopaedics, Leighton General Hospital, Crew, Cheshire, UK. SUBJECTS: A total of 124 patients presenting to A&E with suspected scaphoid fractures between 1 December 1999 and 1 May 2000. MAIN OUTCOME MEASURES: Costs to the hospital budget in ruling out or diagnosing scaphoid fractures in these patients. RESULTS: The cost using an existing protocol that does not routinely involve scanning was pounds 15,554. Projected costs for many (but not all) of the alternative protocols involving routine use of scanning procedures were found to be lower or comparable. CONCLUSIONS: Protocols based on diagnostic scanning for suspected scaphoid fractures can be comparable to a protocol which does not rely on scanning techniques because they can avoid repeated radiographs, clinical appointments and plaster cast changes. However, any such protocol must be tailored on the basis of the workload, and the availability of resources.

Cost-Benefit Analysis↗
PubMed2002

Serological & virological investigation of an outbreak of dengue fever in Gwalior, India.

BACKGROUND & OBJECTIVES: An outbreak of febrile illness occurred between September to November 2001 in Gwalior, Madhya Pradesh affecting individuals mostly in the age group < 30 yr. A total of 312 febrile indoor patients suspected to have dengue infection were investigated. METHODS: The investigation included examination of blood samples from patients for dengue specific IgM and IgG antibodies, isolation of virus in suckling mouse pups and in C(6/36) cell line followed by confirmation and typing through reverse transcriptase-PCR and nested PCR. RESULTS: The serological analysis of the 312 samples indicated 65 per cent positivity of which 21 per cent are of recent infection as indicated by the presence of IgM antibody and 78 per cent are found to be secondary in nature by showing the presence of IgG and/or IgM antibodies. The RT-PCR analysis of patients' sera employing dengue virus group specific conserved amplimer confirmed the etiological agent as dengue complex by showing the characteristic 511 bp amplicons. None of the antibody positive samples were found to be positive by RT-PCR. A total of 13 (6%) samples positive by RT-PCR, were processed for virus isolation in mouse pups and in C(6/36) cells. Of these 9 samples (80%) were confirmed positive for virus isolation as identified by RT-PCR. INTERPRETATION & CONCLUSION: The typing of isolates by nested PCR employing serotype specific amplimer revealed 119 bp amplicon characteristic of dengue virus type-2 and thus confirming the outbreak attributed to dengue virus type-2.

Adult↗
PubMed2001

Role of physiological lung exclusion in difficult lung resections for massive hemoptysis and other problems.

OBJECTIVES: Pulmonary tuberculosis and bronchiectasis are the major causes of massive hemoptysis in developing countries. Lung resection remains the surgical treatment of choice. This may not always be possible and may even be hazardous in some patients due to fibrosis and dense vascular adhesions between the lung and the chest wall. This leads to marked blood loss and control of hilar vessels becomes dangerous. METHODS: A series of 20 cases is described here. Nineteen presented with massive hemoptysis where control of bleeding was obtained by physiological lung exclusion. One patient had traumatic left main bronchus transection not suitable for repair or resection. Physiological lung exclusion was performed by surgical interruption of the bronchus and pulmonary artery of the involved lobe or lung, keeping pulmonary veins intact. RESULTS: Hemoptysis could be controlled in all these patients without any significant morbidity. There was no mortality. There was no postoperative empyema and recurrence of hemoptysis on long-term follow-up. No patient required anatomical lung resection later on. CONCLUSIONS: Physiological lung exclusion is a safe and effective method for control of massive hemoptysis in cases where lung resection is technically hazardous or difficult. This should be kept as an alternative or adjunct to anatomical lung resection.

Adolescent↗
PubMed2001

Evaluation of a dipstick ELISA and a rapid immunochromatographic test for diagnosis of Dengue virus infection.

Here we report standardization of a dipstick enzyme-linked immunosorbent assay (ELISA, Dipstick ELISA) and its comparative evaluation with a commercial Rapid PanBio Immunochromatographic test (IC test) for detection of Dengue (DEN) virus-specific IgM and IgG antibodies in patient sera. Among crude and purified viral antigens prepared from mouse brains or cell cultures, a DEN virus type 2 antigen purified from cell cultures by sucrose density gradient centrifugation was found superior in terms of the signal/ noise (S/N) ratio in the assay system. The sensitivity of detection of the virus by specific IgM antibody was improved by removal of IgG from patient sera prior to testing. The evaluation of the Dipstick ELISA by use of 156 serum samples revealed an overall accordance of 96% and 93% with the IC test in detection of IgM antibodies to DEN viruses (IgM antibodies) and IgG antibodies to DEN viruses (IgG antibodies), respectively. The sensitivity of the Dipstick ELISA and the IC test with reference to the mu-capture ELISA was 83% and 87%, respectively, with a specificity of 98% in both cases. The sensitivity of the Dipstick ELISA with reference to the IC test in detecting IgM and IgG antibodies was 84% and 94%, respectively, and the specificity of the Dipstick ELISA was 98% and 92%, respectively.

Animals↗
PubMed2000

Contralateral hip fractures - can predisposing factors be determined?

A case control study was carried out in the Orthopaedic Department of Bradford Royal Infirmary in an attempt to see if certain medical conditions, which can affect balance and stability, are more common in those who sustain a second proximal femoral fracture. Medical conditions included in the study were: late effects of cerebro-vascular accident, blindness, syncope and collapse, alcoholism, Alzheimer's disease, epilepsy, Parkinsonism, ischaemic heart disease and senile dementia. The study group comprised 53 patients admitted to hospital between 1992 and 1998 with two separate proximal femoral fractures each on a different side. The control group comprised 530 patients selected from a general pool of 2080 proximal femoral fracture patients admitted to hospital during the same period. The control group patients were matched to the study group for age, sex, and time of occurrence of the first fracture. Results show significantly higher association of late effects of cerebro-vascular accident, blindness, syncope and collapse, and Alzheimer's disease with subsequent contralateral proximal femoral fractures. This study supports a causal relationship between the above medical conditions and subsequent contralateral proximal femoral fractures. It may therefore be possible to identify patients who are at risk of returning with a second fracture.

Aged↗
PubMed2000

Study of the relationship of distant SO2 emissions to Dallas-Fort Worth winter haze.

A study was conducted to estimate the changes in wintertime visual air quality in Dallas-Fort Worth (DFW) that might occur due to proposed reductions in SO2 emissions at two steam electric generating plants in eastern Texas, each over 100 km from the city. To provide information for designing subsequent investigations, the haze was characterized broadly during the first year of the study. Meteorological data acquired then demonstrated that, during haze episodes, emissions from only one of the two plants were likely to be transported directly to DFW. Therefore, the second year of the study was centered on just one of the power plants. Air quality was then characterized within the urban area and at rural locations that would be upwind and downwind of the plant during transport to DFW. An instrumented aircraft measured plume dispersion and the air surrounding the plume on selected days. A mathematical model was used to predict the change that would occur in airborne particulate matter concentrations in DFW if SO2 emissions were reduced to reflect the proposed limitations. The contribution of particles in the atmosphere to light extinction was estimated, and simulated photographs were produced to illustrate the visibility changes. The study concluded that the proposed emission reductions would, at most, subtly change perceived wintertime visibility.

Air Pollution↗
PubMed2000

Concentration and composition of atmospheric aerosols from the 1995 SEAVS experiment and a review of the closure between chemical and gravimetric measurements.

We summarize the results from the various measurements and the inter-sampler comparisons from Southeastern Aerosol and Visibility Study (SEAVS), a study with one of its objectives to test for closure among chemical, gravimetric and optical measurements of atmospheric aerosol particles. Sulfate and organics are the dominant components of the SEAVS fine particles (nominally, particles with aerodynamic diameter < or = 2.5 microns) but between 28 and 42% (range over various samplers) of the gravimetrically measured total fine particle concentration is unidentified by the chemical measurements. Estimates of water associated with inorganic components and measurement imprecision do not totally explain the observed difference between gravimetric and chemical measurements. We examine the theoretical and empirical basis for assumptions commonly made in the published literature to extrapolate total fine particle concentration on the basis of chemical measurements of ions, carbon and elements. We then explore the more general question of closure using the SEAVS data as well as data from other, similar studies reported in the literature. In so combining the SEAVS measurements with other similar studies, we find a strong association between organic carbon and the unidentified component, that is, the fraction of the total fine particle concentration not identified by chemical measurements. We offer several tenable hypotheses for the relationship between the organic and unidentified components that deserve to be tested in future work. Specifically, we hypothesize that (1) errors in the sampling and analysis of organic carbon; (2) estimates of organic mass from measurements of organic carbon; and/or (3) water absorption by organics may all contribute to the observed relationship.

Aerosols↗
PubMed2000

Modeling of potential power plant plume impacts on Dallas-Fort Worth visibility.

During wintertime, haze episodes occur in the Dallas-Ft. Worth (DFW) urban area. Such episodes are characterized by substantial light scattering by particles and relatively low absorption, leading to so-called "white haze." The objective of this work was to assess whether reductions in the emissions of SO2 from specific coal-fired power plants located over 100 km from DFW could lead to a discernible change in the DFW white haze. To that end, the transport, dispersion, deposition, and chemistry of the plume of a major power plant were simulated using a reactive plume model (ROME). The realism of the plume model simulations was tested by comparing model calculations of plume concentrations with aircraft data of SF6 tracer concentrations and ozone concentrations. A second-order closure dispersion algorithm was shown to perform better than a first-order closure algorithm and the empirical Pasquill-Gifford-Turner algorithm. For plume impact assessment, three actual scenarios were simulated, two with clear-sky conditions and one with the presence of fog prior to the haze. The largest amount of sulfate formation was obtained for the fog episode. Therefore, a hypothetical scenario was constructed using the meteorological conditions of the fog episode with input data values adjusted to be more conducive to sulfate formation. The results of the simulations suggest that reductions in the power plant emissions lead to less than proportional reductions in sulfate concentrations in DFW for the fog scenario. Calculations of the associated effects on light scattering using Mie theory suggest that reduction in total (plume + ambient) light extinction of less than 13% would be obtained with a 44% reduction in emissions of SO2 from the modeled power plant.

Aerosols↗
PubMed2000

Transition metal-catalyzed oxidation of ascorbate in human cataract extracts: possible role of advanced glycation end products.

PURPOSE: With age, human lens crystallins become more pigmented, oxidized, modified by ascorbate oxidation and advanced glycation end products (AGEs), and bind copper. The hypothesis was tested that the major AGE and ascorbylation product in the human lens, N(epsilon)-carboxymethyl-L-lysine (CML), has an EDTA-like structure, which may predispose it to bind redox active copper. METHODS: Young, old, and cataractous human lens protein fractions were glycated with ascorbic acid and tested for their ability to bind Cu(II) by atomic absorption spectroscopy and oxidize (14C1)-ascorbate by radiometric thin-layer chromatography method. AGEs were assayed by high-performance liquid chromatography (HPLC). CML-rich proteins were immunoprecipitated from young, old, and cataractous crystallins using affinity-purified CML antibody and tested for their ability to oxidize ascorbate and generate hydroxyl radicals in the presence of H2O2 using 5,5'-dimethyl-1-pyrroline-N-oxide (DMPO) spin-trap and EPR spectroscopy. RESULTS: Ascorbate oxidizing activity at 24 hours of native crystallins was significantly increased in both the water soluble (WS; P < 0.001) and insoluble (WIS; P < 0.05) fractions from cataractous and normal lenses. The chelator DTPA completely prevented oxidation up to 24 hours of incubation but less effectively thereafter. Mean endogenous Cu content in pooled young, old, and cataract fractions increased from 0.016 to 0.026 nmol/mg protein, respectively, in WS (P < 0.05) and WIS (P < 0.001) fractions, and Cu(II) binding was 20% to 30% increased in cataractous versus old and young lenses in WS (P < 0.01) and WIS (P < 0.001) fractions. Mean levels of the AGEs, CML, and pentosidine were markedly elevated in WS and WIS fractions from cataractous versus old or young crystallins (20% to severalfold, P < 0.05 to P < 0.001). In a separate experiment, protein-bound Fe was not elevated. Crystallins ascorbylated in vitro showed an increase in CML as well as Cu(II) binding. CML-rich proteins (immunoprecipitated from cataractous lenses) oxidized ascorbate approximately 4 times faster than similar proteins from young and old normal lenses (P < 0.01) and generated hydroxyl radicals in the presence of H2O2 and DMPO. CONCLUSIONS: The association between CML formation, copper binding, and generation of free radicals by cataractous lens crystallins can be duplicated by ascorbylation in vitro. These effects are only in part attributable to CML itself, and other modifications (AGEs, conformational changes) may participate in the process. A vicious cycle between AGE formation, lipoxidation, and metal binding may exist in the aging lens, suggesting that chelation therapy could be beneficial in delaying cataractogenesis.

Ascorbic Acid↗
PubMed1999

Protein aging by carboxymethylation of lysines generates sites for divalent metal and redox active copper binding: relevance to diseases of glycoxidative stress.

Aging and age-related diseases are associated with the production of reactive oxygen species which modify lipids, proteins and DNA. Here we hypothesized the glyco- and lipoxidation product N(epsilon)-(carboxymethyl)lysine (CML) in proteins should bind divalent and redox active transition metal binding. CML-rich poly-L-lysine and bovine serum albumin (BSA) were chemically prepared and found to bind non-dialyzable Cu(2+), Zn(2+) and Ca(2+). CML-BSA-copper complexes oxidized ascorbate and depolymerized protein in the presence of H(2)O(2). CML-rich tail tendons implanted for 25 days into the peritoneal cavity of diabetic rats had a 150% increase in copper content and oxidized ascorbate three times faster than controls. CML-rich proteins immunoprecipitated from serum of uremic patients oxidized four times more ascorbate than control and generated spin adducts of DMPO in the presence of H(2)O(2). The chelator DTPA suppressed ascorbate oxidation thereby implicating transition metals in the process. In aging and disease, CML accumulation may result in a deleterious vicious cycle since CML formation itself is catalyzed by lipoxidation and glycoxidation.

Animals↗
PubMed1999

Canine external carotid vasoconstriction to methysergide, ergotamine and dihydroergotamine: role of 5-HT1B/1D receptors and alpha2-adrenoceptors.

The antimigraine drugs methysergide, ergotamine and dihydroergotamine (DHE) produce selective vasoconstriction in the external carotid bed of vagosympathectomized dogs anaesthetized with pentobarbital and artificially respired, but the receptors involved have not yet been completely characterized. Since the above drugs display affinity for several binding sites, including alpha-adrenoceptors and several 5-HT1 and 5-HT2 receptor subtypes, this study has analysed the mechanisms involved in the above responses. Intracarotid (i.c.) infusions during 1 min of methysergide (31-310 microg min(-1)), ergotamine (0.56-5.6 microg min(-1)) or DHE (5.6-31 microg min(-1)) dose-dependently reduced external carotid blood flow (ECBF) by up to 46+/-4, 37+/-4 and 49+/-5%, respectively. Blood pressure and heart rate remained unchanged. The reductions in ECBF by methysergide were abolished and even reversed to increases in animals pre-treated with GR127935 (10 microg kg(-1), i.v.). The reductions in ECBF by ergotamine and DHE remained unchanged in animals pre-treated (i.v.) with prazosin (300 microg kg(-1)), but were partly antagonized in animals pre-treated with either GR127935 (10 or 30 microg kg(-1)) or yohimbine (1000 microg kg(-1)). Pre-treatment with a combination of GR127935 (30 microg kg(-1)) and yohimbine (1000 microg kg(-1)) abolished the responses to both ergotamine and DHE. The above doses of antagonists were shown to produce selective antagonism at their respective receptors. These results suggest that the external carotid vasoconstrictor responses to methysergide primarily involve 5-HT1B/1D receptors, whereas those to ergotamine and DHE are mediated by 5-HT1B/1D receptors as well as alpha2-adrenoceptors.

Adrenergic alpha-1 Receptor Antagonists↗