A large-volume sputum dry storage and transportation device for molecular and culture-based diagnosis of tuberculosis.

Technologies for preservation of specimens in the absence of cold chains are essential for optimum utilization of existing laboratory services in the developing world. We present a prototype called specimen transportation tube (SPECTRA-tube) for the collection, exposure-free drying, ambient transportation, and liquid state recovery of large-volume (>1 mL) specimens. Specimens introduced into the SPECTRA-tube are dried in glass fiber membranes, which are critical for efficient liquid-state sample recovery by rehydration and centrifugation. SPECTRA-tube is demonstrated for the dry storage of sputum for tuberculosis detection. Mycobacterium smegmatis (Msm)-spiked mock sputum dried in a native Standard 17 glass fiber was stable for molecular testing after 10 day storage at 45 °C and for culture testing after 10- and 5-day storage at 37 °C and 45 °C, respectively. Compatibility with human sputum storage was demonstrated by dry storing 1.2 mL Mycobacterium bovis-spiked human sputum in a SPECTRA-tube for 5 days at room temperature. We have thus demonstrated the first workflow for dry storage of sputum followed by molecular and culture testing. Compared to existing specimen dry storage technologies, SPECTRA-tube significantly increases the volume of liquid specimens that can be transported in the dry state and enables the recovery of the entire sample in the liquid state, rendering it compatible with conventional downstream analysis methods.

Antibiotic resistance: A cross-sectional study on knowledge, attitude, and practices among veterinarians of Haryana state in India.

AIM: The current study aimed to assess the knowledge, attitude, and practices pertaining to antibiotic usage among the field veterinarians who serve as nodal officers playing a crucial role in disseminating knowledge to the farmers regarding livestock management practices in India. MATERIALS AND METHODS: A pilot study was conducted in which 106 of the 173 field veterinarians of Haryana, India, agreed to contribute through their valuable participation in the study. The collected data were critically analyzed by simple descriptive statistics, and the responses were ranked using Garrett's ranking method. RESULTS: Our study found that most of the clinicians were aware of the fundamental clinical aspects of antibiotic resistance (AR), i.e., the general causes and transmission of resistance, response during treatment failure, and safe disposal of hospital waste. Further, implementation of "antibiotic stewardship" (rational/responsible use of antibiotics) and interruption of AR transmission by means of cross-kingdom pathogens are two ways to restrict the spread of resistant pathogens which were not in the clinical purview of majority of the clinicians. This highlights a lack of awareness and scope of improving clinician's knowledge pertaining to AR. Moreover, we got to know the methodology adopted by farmers for disposal of infected milk from diseased udders as well as their attitude toward diseased and unproductive animals. CONCLUSION: This study provides snippets of the current animal husbandry practices prevalent at the field level which would assist to plug in the gaps of knowledge regarding AR among the veterinarians as well as the general public and serve to reduce its deleterious impacts in Indian animal farming as well as in the world through the concept of "One World, One Health."

Evidences for diabetes and insulin mimetic activity of medicinal plants: Present status and future prospects.

Diabetes mellitus (DM) is a considerable systemic metabolic disorder to exhibit various metabolic and cardiovascular disorders, mainly hyperglycemia. The global projected estimate of diabetes in 2030 will be about 439 million adults, out of which 300 million expected are of type-2 diabetes mellitus (T2DM). The present knowledge revealed responsible factors, occurrence and mechanism of these factors involved in the DM diseases. Hence, the aim of this review is to address and summarize the causes, plant resources, importance, present status and future programmes for diabetes control. The present review answers the contemporary present questions raised in the scientific field on DM. Two major problems are explained in detail about the autoimmune attack or dysfunction of ß-cell and insulin resistance involved for Type 1 and Type 2 DM, respectively. Though there are various approaches to reduce the ill effects of diabetes and its secondary complications, many preferred herbal formulations due to lesser side effects and low cost. For this reason still it is getting increased attention in searching antidiabetic medicinal plants for hot research and to develop targeted medicine. Recurrence of islet autoimmunity lesson from pancreatic islet cell transplantation to cure T1D was outlined. With these highlights, the review summarizes the current knowledge on diabetes occurrence, factors (environmental and genetics), and types (I, II, gestation, and secondary DM), antidiabetic plants, sources for insulin mimetic plant principle compounds and their target mechanism with current and future trusted research areas for controlling of DM.

Novel T-C@AgNPs mediated biocidal mechanism against biofilm associated methicillin-resistant Staphylococcus aureus (Bap-MRSA) 090, cytotoxicity and its molecular docking studies.

Staphylococcus aureus is a commonly found pathogen that can cause food-spoilage and life threatening infections. However, the potential molecular effects of natural active thymol molecules and chitosan silver nanoparticles (C@AgNPs) in bacteria remain unclear. This gap in the literature has prompted us to study the effects of thymol loaded chitosan silver nanoparticles (T-C@AgNPs) against biofilm associated proteins in methicillin-resistant S. aureus (Bap-MRSA) 090 and also their toxicity, anti-cancer activity, and validation of their in silico molecular docking. The results showed excellent antibacterial activity of T-C@AgNPs against Bap-MRSA 090, having a minimum inhibitory concentration of 100 µg mL-1 and a 10.08 ± 0.06 mm zone of inhibition (ZOI). The cyclic voltammogram (CV) analysis clearly showed pore forming of T-C@AgNPs at 300 µg mL-1 concentration, and evidence of the interruption of the electron transport chain was clearly seen. The 200 µg mL-1 concentration exhibited a 52.60 ± 0.25% anti-biofilm property by T-C@AgNPs against Bap-MRSA 090. The T-C@AgNPs showed no toxicity to peripheral blood mononuclear cells (PBMC) (IC50 = 221 ± 0.71 µg mL-1) compared to the control, and anti-cancer activity against human triple negative breast cancer cell line (MDA-MB-231) (IC50 110 ± 1.0 µg mL-1) compared to the standard drug Doxorubicin (IC50 = 19 ± 1.0). The excellent properties of T-C@AgNPs were validated by in silico molecular docking studies and showed best match scoring to target proteins compared to standards. These excellent properties of T-C@AgNPs highlight for the first time its pharmacology and potential in medicinal drug development applications for future research.

Synthesis of 4-Isoxazolines through Gold(I)-Catalyzed Cyclization of Propargylic N-Hydroxylamines.

New catalytic methods for the synthesis of 4-isoxazolines have been developed via catalytic intramolecular cyclizations of propargylic N-hydroxylamines. The reactions proceed rapidly in less than 1 h at room temperature in the presence of 5 mol % (PPh3)AuCl/5 mol % AgOTf or 5 mol % (PPh3)AuNTf2. This process features an efficient route to 4-isoxazolines with high yields, short reaction times, and mild reaction conditions.

Immunoreactivity and trypsin sensitivity of recombinant virus-like particles of foot-and-mouth disease virus.

Foot-and-mouth disease (FMD) is an important infection affecting the health and productivity of cloven-hoofed livestock. Development of improved vaccines and diagnostic reagents is being explored to facilitate the disease control. There is an emerging interest in virus-like particles (VLPs), as their constituent structural proteins are the major immunogens. The VLPs are similar to natural virus particles but lack viral nucleic acid. The objective of the present study was to express the VLPs of FMD virus (FMDV) serotype Asia-1 (IND 63/72), using baculovirus system and characterize them for antigenic structure. The VLPs expressed in insect cells showed immunoreactivity similar to inactivated cell culture FMDV. Further they possess similar sensitivity to trypsin as the inactivated cell culture FMDV, suggesting that trypsin-sensitive antigenic sites could be similarly arranged. Our findings suggest that the FMD VLPs have similar antigenic conformational feature like the wild type virus, thus supporting their utility in development of non-infectious FMD vaccines and/or diagnostic assays.

Hydrophobic interactions in donor-disulphide-acceptor (DSSA) probes looking beyond fluorescence resonance energy transfer theory.

Donor-linker-acceptor (DSSA) is a concept in fluorescence chemistry with acceptor being a fluorescent compound (FRET) or quencher. The DSSA probes used to measure thiol levels in vitro and in vivo. The reduction potential of these dyes are in the range of -0.60 V, much lower than the best thiol reductant reported in literature, the DTT (-0.33 V). DSSA disulphide having an unusually low reduction potential compared to the typical thiol reductants is a puzzle. Secondly, DSSA probes have a cyclized rhodamine ring as acceptor which does not have any spectral overlap with fluorescein, but quenches its absorbance and fluorescence. To understand the structural features of DSSA probes, we have synthesized DSSANa and DSSAOr. The calculated reduction potential of these dyes suggest that DSSA probes have an alternate mechanism from the FRET based quenching, namely hydrophobic interaction or dye to dye quenching. The standard reduction potential change with increasing complexity and steric hindrance of the molecule is small, suggesting that ultra- low Eo' has no contribution from the disulphide linker and is based on structural interactions between fluorescein and cyclized rhodamine. Our results help to understand the DSSA probe quenching mechanism and provide ways to design fluorescent probes.

Design of multiligand inhibitors for the swine flu H1N1 neuraminidase binding site.

Viral neuraminidase inhibitors such as oseltamivir and zanamivir prevent early virus multiplication by blocking sialic acid cleavage on host cells. These drugs are effective for the treatment of a variety of influenza subtypes, including swine flu (H1N1). The binding site for these drugs is well established and they were designed based on computational docking studies. We show here that some common natural products have moderate inhibitory activity for H1N1 neuraminidase under docking studies. Significantly, docking studies using AutoDock for biligand and triligand forms of these compounds (camphor, menthol, and methyl salicylate linked via methylene bridges) indicate that they may bind in combination with high affinity to the H1N1 neuraminidase active site. These results also indicate that chemically linked biligands and triligands of these natural products could provide a new class of drug leads for the prevention and treatment of influenza. This study also highlights the need for a multiligand docking algorithm to understand better the mode of action of natural products, wherein multiple active ingredients are present.

Utility of concentration method by modified bleach technique for the demonstration of acid-fast bacilli in the diagnosis of tuberculous lymphadenopathy.

BACKGROUND: Microscopy detection of acid fast bacilli (AFB) by Ziehl-Neelsen (ZN) method has many advantages when it comes to speed and feasibility though it has a low sensitivity. If the sensitivity could be improved, it has the potential to become an even more valuable tool for detection of AFB. OBJECTIVES: To evaluate the efficacy of bleach concentration method in the cytodiagnosis of tuberculous lymphadenitis in comparison with routine Ziehl-Neelsen method and to compare the positivity in various cytomorphological categories. MATERIALS AND METHODS: A total of 112 cases of tuberculous lymphadenitis diagnosed by fine needle aspiration cytology (FNAC) were categorized into six cytomorphological patterns. The acid-fast bacilli positivity by routine staining was correlated with modified bleach methods of ZN staining. Sensitivity of routine ZN and modified bleach concentration was compared. RESULTS: The classic cytomorphological pattern of tuberculosis of epithelioid granulomas, langhans giant cells and caseous necrosis was seen in 37.5% of cases. Routine ZN staining detected AFB in 12.5% of cases and the modified bleach method in 60.7%. Modified bleach method showed AFB positivity in additional 54 cases where routine AFB staining was negative. The modified bleach method showed AFB positivity in all cases where routine ZN staining was positive. CONCLUSION: The modified bleach method was more sensitive and safer than routine ZN staining. As the background was clear, the bacilli were easily visible and the screening time was shorter.

Bio-analytical method development and validation of Rasagiline by high performance liquid chromatography tandem mass spectrometry detection and its application to pharmacokinetic study.

The most suitable bio-analytical method based on liquid-liquid extraction has been developed and validated for quantification of Rasagiline in human plasma. Rasagiline-13C3 mesylate was used as an internal standard for Rasagiline. Zorbax Eclipse Plus C18 (2.1 mm×50 mm, 3.5 µm) column provided chromatographic separation of analyte followed by detection with mass spectrometry. The method involved simple isocratic chromatographic condition and mass spectrometric detection in the positive ionization mode using an API-4000 system. The total run time was 3.0 min. The proposed method has been validated with the linear range of 5-12000 pg/mL for Rasagiline. The intra-run and inter-run precision values were within 1.3%-2.9% and 1.6%-2.2% respectively for Rasagiline. The overall recovery for Rasagiline and Rasagiline-13C3 mesylate analog was 96.9% and 96.7% respectively. This validated method was successfully applied to the bioequivalence and pharmacokinetic study of human volunteers under fasting condition.

Noise induced hearing loss (NIHL) in saw mill and printing press workers in Akluj Town of Solapur district.

Noise pollution and allied health problems are seen at all ages worldwide. This is due to increase in mechanization in industries, transportation as well as home appliances produce high level of noise. Noise Induced Hearing Loss (NIHL) is common health complaint found in industrial workers. The present research work reveals the NIHL problem in workers related to saw mill and printing press in Akluj town, Solapur district of Maharashatra state. For the present study measurement of noise levels are done in saw mills and printing presses. The standard noise level value is calculated in respective locations. Noise Induced Hearing Loss (NIHL) among the workers is measured by an Audiometer. 420 workers, viz. 250 from ten saw mills and 170 from ten printing presses respectively were investigated to find out NIHL. The average noise levels in printing press and saw mill are 90.2 dB and 79.3 dB respectively. Whereas standard noise level (Leq) in above locations are 101.4 dB and 98.7 dB respectively. The average NIHL are found in 28% saw mill workers and 13% in printing press workers. It is reported that medicine treatments are not very useful for curing NIHL. Today's available tool which prevents NIHL is ear plugs and ear muffs, which are suitable for preventing NIHL problems in industrial workers.

A new method to generate an almost-diagonal matrix in the boundary integral equation formulation.

In this work, a new numerical procedure is developed to generate an almost-diagonal matrix for the solution of boundary integral equation formulation dealing with acoustic scattering problems. The major drawback of the traditional boundary integral equation procedure resulting in a dense system matrix is eliminated in this new procedure by grouping the basis functions into a cluster. The geometry of the structure is modeled by planar triangles and the basis functions are defined on the nodes. By doing so, one can get a benefit of a smaller size matrix to begin with. Furthermore, by grouping these node-based basis functions into a cluster, an almost-diagonal matrix is generated. Thus, the solution procedure developed in this work may be utilized for very large scattering problems since the required computer resources are very low. The solution procedure developed in this work is validated for the scattering cross section of the simple shapes with the closed form solutions wherever available and with the other numerical solution procedures.

In search of a pediatric cardiac surgeon's 'Holy Grail': the ideal pulmonary conduit.

The limited lifespan of all currently available conduits leads to repeat operations and interventional procedures in many children. Each reoperation entails considerable risk to life, expenditure and compromised quality of life as the conduit degenerates. The ideal conduit should be available freely, inexpensive, require no anticoagulation, be resistant to infection, free from thromboembolism, have no gradients or regurgitation and have unlimited durability. This review explores various options as surgeons and researchers endeavor to develop the ideal conduit--which will fulfill all of the above-mentioned criteria. Various currently available conduits are analyzed. Special emphasis is given to tissue-engineered valves and percutaneous valve implantations.

A novel nonphagocytic mechanism of erythrocyte destruction involving direct cell-mediated cytotoxicity.

The number of erythrocytes fell when co-cultured with cell preparations derived from mouse spleen, thymus, bone marrow, or peritoneal exudate (PE) cells. Erythrocyte-depletion activities (EDA) of different leukocyte preparations were in the order PE > spleen > thymus > bone marrow. Adherent, nonadherent, T-depleted, and T-enriched cell subpopulations had comparable EDA. Spleen cells from athymic nude mice, however, lacked significant EDA. In addition, EDA was boosted by Concanavalin A (Con A) but not by lipopolysaccharide, indicating that T cells may play a crucial role in inducing EDA in spleen cells. Paraformaldehyde-fixed spleen or PE cells, as well as membrane preparations isolated from spleen cells, efficiently lysed erythrocytes. Erythrocyte ghost membranes inhibited erythrocyte lysis by control or paraformaldehyde-fixed spleen cells. Treatment with hamster anti-mouse Fas or anti-mouse tumor necrosis factor receptor (TNFR) antibody could opsonize erythrocytes for faster depletion by spleen cells, suggesting an expression of Fas and TNFR on erythrocytes. TNF alpha could lyse erythrocytes in a dose-dependent fashion. Additionally, enhanced spleen cell EDA induced in response to succenyl Con-A could be blocked by anti-TNF alpha antibodies. Our results provide evidence for a direct cell-mediated cytotoxicity (CMC) of erythrocytes by leukocytes. A role of molecules of Fas and the TNF family in CMC of erythrocytes has also been suggested. Further work is needed to understand if, and to what extent, CMC of erythrocytes contributes to erythrocyte destruction in vivo and to determine its patho-physiological significance.

Salvage and reconstruction of complex post-extirpative knee wounds: a successful surgical protocol.

Six patients with massive post-extirpative wounds of the knee, all of whom had received adjuvant therapy, underwent wound closure and limb salvage via free-tissue transfer in all but one case. No instance of delayed wound healing requiring surgery occurred among the patients reconstructed by microsurgical tissue transfer. Five of the six patients are presently ambulating on the salvaged extremity. A protocol for management of these complex wounds is presented.

Cephalic vein in salvage microsurgical reconstruction in the head and neck.

Free tissue transfers have significantly improved the outcome of major head and neck reconstruction. In some situations of microvascular tissue transfer, adequate recipient veins are not available for flap venous outflow. This may result from a variety of reasons, including prior radical neck dissection, radiation therapy, and inflammatory changes from severe trauma. We report our experience using cephalic vein transposition in 11 such patients with unavailable local veins for free flap reconstruction. In all cases the cephalic vein provided reliable and adequate outflow for the tissue transfers. Our experience suggests that the cephalic vein transposition offers certain advantages, obviating the use of vein grafts: 1. It requires only one venous anastomosis; 2. A long pedicle can be harvested to reach the mid-face or contralateral neck without undue tension; 3. The cephalic-subclavian system is a high-flow, low-pressure system; 4. This vein is located outside ablative surgical field, or radiated tissue, and therefore undamaged; 5. The vein calibre is well suited for microsurgical anastomosis.

Evidence for cell-surface association between fusin and the CD4-gp120 complex in human cell lines.

Accessory cell-surface molecules involved in the entry of human immunodeficiency virus-type 1 into cells have recently been identified and shown to belong to the family of chemokine receptors. Treatment of human cell lines with soluble monomeric gp120 at 37 degrees C induced an association between the surface CD4-gp120 complex and a 45-kilodalton protein, which can be down-modulated by the phorbol ester phorbol 12-myristate 13-acetate. The three proteins were coprecipitated from the cell membranes with antibodies to CD4 or to gp120. The 45-kilodalton protein comigrated with fusin on sodium dodecyl sulfate gels and reacted with rabbit antisera to fusin in protein immunoblots. No 45-kilodalton protein could be coprecipitated from similarly treated nonhuman cells. However, infection of 3T3.CD4.401 cells with vaccinia-fusin recombinant virus (vCBYF1), followed by gp120 treatment, resulted in coprecipitation of fusin and CD4.401 molecules from their membranes. Together these data provide evidence for physical association between fusin and the CD4-gp120 complex on cell membranes.

Surgical treatment of complications 45 years after extraperiosteal pneumonolysis and plombage using acrylic resin balls for cavitary pulmonary tuberculosis.

An infected axillary sinus tract discharged balls made of an acrylic resin consisting essentially of polymerized methyl methacrylate (Lucite) 45 years following performance of an extraperiosteal pneumonolysis and Lucite ball plombage for collapse therapy of right upper lobe cavitary tuberculosis. Surgical extraction of the balls was performed, followed by a partial decortication of the lung and intrathoracic transposition of a pectoralis major muscle flap to fill the residual pleural space. Primary healing was attained, and the patient is well 1 1/2 years after surgery.