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Background: Methicillin-resistant Staphylococcus aureus (MRSA) is a cause of increased morbidity and mortality in debilitated and immunocompromised patients., Aims and Objectives: To study the prevalence of MRSA among dermatology inpatients and elucidate its predisposing factors and antibiotic sensitivity pattern., Materials and Methods: This prospective study involved all the inpatients in the dermatology ward with suspected bacterial infections, both primary and secondary. Gram's stain, pus culture, and antibiotic sensitivity were the main tests done., Results: There were 61 patients in this study, with 36 males (59%) and 25 females (41%). The age group maximally affected was 41-50 years (21.6%). Vesiculobullous disorders (22, 36.1%) and eczemas (16, 26.2%) were the commonest dermatoses with secondary infection. Extremities (72.1%) and the trunk (26.2%) were the commonest sites involved. Diabetes mellitus (DM) was the commonest systemic illness (11.5%). Staphylococcus aureus was the commonest isolate, grown in 52 patients (85.2%) of whom 16 (30.80%) were MRSA positive. MRSA constituted 22.2% of the total bacterial isolates. All the MRSA isolates (100%) were resistant to cloxacillin and oxacillin, while all the isolates (100%) were sensitive to vancomycin and linezolid., Conclusions: The prevalence of MRSA in this study was high. It was most commonly seen in patients with vesiculobullous disorders. A high sensitivity of MRSA was found to amikacin. There was no relation between prolonged antibiotic therapy and MRSA in this study.

Hospital acquired infections are prevalent in many hospitals and contaminated touch surfaces are a route of transmission. To find a solution for this, copper and copper oxide thin films were deposited via aerosol assisted chemical vapour deposition using copper nitrate as the precursor and the films were characterised by a range of techniques including powder X-ray diffraction and scanning electron microscopy. The antimicrobial activity of the deposited copper and copper oxide films were investigated against Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli). A 5-log 10 reduction in the viable counts of E. coli was observed on the copper thin films after 30 minutes while a 2-log 10 reduction was observed on copper oxide films after 1 hour. In the case of S. aureus both copper and copper oxide films exhibited 4-log 10 reduction after 1 hour. The high antimicrobial efficacy of the Cu 2 O films, approaching that of the pure copper films, suggests that oxide formation on copper objects should not significantly impair their antimicrobial activity.

Introduction: Obesity is a condition in which excess body fat accumulates, which leads to various adverse effects on health, particularly cardiovascular diseases (CVDs), which reduce life expectancy and/or increase health problems. Fast food consumption is one of the factors which have been reported as a cause of obesity. Body mass index (BMI) is used to assess obesity and overweight, which can be calculated by using the formula, weight in kg, divided by square of height in metres., Aim: This study focused on the relationship of body mass index with fast food consumption, associated soft drink consumption and physical activity., Methods: Descriptive cross-sectional study was conducted in Department of Biochemistry, SBKS MI and RC, and Sumandeep Vidyapeeth. This study was approved by the ethical review board .One hundred and forty seven medical students from 1(st) year MBBS course were included in this study. Self-structured questionnaire was used, which contained several data like information on age, height, weight, education level. The formula used for calculating BMI was, weight in kg, divided by square of height in metres (Kg/m(2))., Results: In our study, out of 147 students, a total of 138 students (more than 90%) used to have fast food. Among these, a total of 47 students (34.05%) were pre-obese and obese. Out of 147 students, 87 students (59.18%) were in normal weight range, while 13 (8.84%) students were underweight., Statistical Analysis: Data was compiled in an Excel worksheet and it was analyzed for percentages and proportions. Chi-square and Pearson's correlation test were also applied wherever they were applicable and Alpha error was set at a 5% level., Conclusion: In our study, a significant relationship was found between BMI and fast food consumption, less physical activity, and intake of soft drinks.

Background: Antibiotic therapy can select for small colony variants of Staphylococcus aureus that are more resistant to antibiotics and can result in persistent infections, necessitating the development of more effective antimicrobial strategies to combat small colony variant infections. Photodynamic therapy is an alternative treatment approach which utilises light in combination with a light-activated antimicrobial agent to kill bacteria via a non-specific mechanism of action. In this study, we investigated whether the combination of 665 nm laser light and the light-activated antimicrobial agent methylene blue was able to successfully kill S. aureus small colony variants. S. aureus and isogenic stable small colony variant were exposed to varying doses (1.93 to 9.65 J/cm2) of 665 nm laser light in the presence of varying concentrations (1 to 20 μM) of methylene blue., Results: The combination of 665 nm laser light and methylene blue was found to be an effective strategy for the killing of small colony variants. At the highest light dose (9.65 J/cm2) and methylene blue concentration (20 μM) tested, the number of viable bacteria decreased by approximately 6.9 log10 for the wild type and approximately 5 log10 for the small colony variant., Conclusions: These results suggest that photodynamic therapy has potential for use in the treatment of superficial infections caused by small colony variants of S. aureus and supports further research in this field.

Introduction: Health professionals need to develop analytic and diagnostic thinking skills and not just a mere accumulation of large amount of facts. Hence, Case Based Learning (CBL) has been used in the medical curriculum for this reason, so that the students are exposed to the real medical problems, which helps them in develop analysing abilities. This also helps them in interpreting and solving the problems and in the course of doing this, they develop interest. In addition to didactic lectures, CBL was used as a learning method., Methods: This study was conducted in the Department of Biochemistry, S.B.K.S.M.I and R.C, Sumandeep Vidyapeeth ,Piparia, Gujarat, India. A group of 100 students were selected and they were divided into two groups as the control group and the study group. A total of 50 students were introduced to case based learning, which formed the study group and 50 students who attended didactic lectures formed the control group., Results: A very significant improvement (p<0.0001) was observed among the students after the CBL sessions and they were also motivated by these sessions. A 4 point Likert scale questionnaire which contained 8 questions was administered to the students, to know their perception on the usefulness of the CBL. 98% of the students reported that they found the CBL sessions to be an interesting method of gaining knowledge. 84% of them felt that they exposed them to an experience of logical application of the knowledge which was gained in cracking cases, which would be of great help in the future also., Conclusion: Case Based Learning (CBL) was used and it is effective in the medical curriculum for a better understanding of Biochemistry among the medical students.

Background: Histoid leprosy (HL) is a rare variant of lepromatous leprosy presenting with well-defined smooth shiny papules and nodules. This can mimic many other dermatoses and can be missed clinically., Methods: This is a retrospective 10-year study. Data were obtained and analyzed from case records of the urban leprosy clinic at the Government Medical College, Trivandrum, Kerala (India)., Results: There were 17 cases of HL in this study. The male/female ratio was 16 : 1 and the 41-50-year-old age group accounted for the maximum number of cases (six, 35.29%); 11 cases (64.70%) were de novo cases of HL. Papules were the commonest skin lesions encountered (17, 100%), while the upper and lower limbs and trunk were the most common sites involved (11, 64.70%). Earlobe infiltration was seen in 10 cases (58.82%). Lepra reactions were seen in four (23.52%) cases. Deformity/disability occurred only in two cases (11.76%). The mean morphological index was 30.58%, while the mean bacteriological index was 5.17. Epidermal atrophy and a well-circumscribed dermal area packed with spindle-shaped histiocytes and positive for acid-fast bacilli were the main histopathological features seen., Conclusions: The majority of cases in this study were de novo. HL can occur even when there is no history of inadequate or irregular treatment. Papules rather than nodules were the most common skin lesions seen. Earlobe infiltration was seen in the majority of cases unlike that reported in the literature. Lepra reactions are very rare in HL., (© 2013 The International Society of Dermatology.)

This study presents a novel way of enhancing plant growth through the use of a non-petroleum based product. We report here that exposing either roots or seeds of multicellular plants to extremely low concentrations of dissolved hydrogen sulfide at any stage of life causes statistically significant increases in biomass including higher fruit yield. Individual cells in treated plants were smaller (~13%) than those of controls. Germination success and seedling size increased in, bean, corn, wheat, and pea seeds while time to germination decreases. These findings indicated an important role of H2S as a signaling molecule that can increase the growth rate of all species yet tested. The increased crop yields reported here has the potential to effect the world's agricultural output.

Background: Gold nanoparticles (AuNP) are extensively used as biophysical tools in the area of medicine and technology due to their distinct properties. However, vivid understanding of the consequences of biomolecule-nanomaterial interactions is still lacking. In this context, we explore the affect of conjugation of Gαi1 subunit (of heterotrimeric G-proteins) to AuNP and examine its consequences. We consider two bio-conjugation strategies covalent and non-covalent binding., Results: Affinity of the AuNP to the Gαi1 is 7.58 × 10 12 M-1. AuNP conjugated Gαi1 exhibits altered kinetics of activation, non-covalent bio-conjugates displays retarded kinetics, up to 0.88 fold when GTPγS was used as ligand, of protein activation contrary to covalent conjugates which accelerates it to ~ 5 fold. Conjugation influence intrinsic Gαi1 GTPase function in conflicting modes. Non-covalent conjugation inhibits GTPase function (decrease in activity upto 0.8 fold) whilst covalent conjugation drastically accelerates it (12 fold increase in activity). Altered basal nucleotide uptake in both types of conjugates and GTPase function in non-covalent conjugate are almost comparable except for GTPase property of covalent conjugate. The effect is despite the fact that conjugation does not change global conformation of the protein., Conclusion: These findings provide clear evidence that nanoparticles, in addition to 'passive interaction' with protein (biomolecule), can interact "actively" with biomolecule and modify its function. This concept should be considered while engineering nanoparticle based delivery systems in medicine.

Rodent models of absence seizures are used to investigate the network properties and regulatory mechanisms of the seizure's generalized spike and wave discharge (SWD). As rats age, SWDs occur more frequently, suggesting aging-related changes in the regulation of the corticothalamic mechanisms generating the SWD. We hypothesized that brain resetting mechanisms - how the brain "resets" itself to a more normal functional state following a transient period of abnormal function, e.g., a SWD - are impaired in aged animals and that brain infarction would further affect these resetting mechanisms. The main objective of this study was to determine the effects of aging, infarction, and their potential interaction on the resetting of EEG dynamics assessed by quantitative EEG (qEEG) measures of linear (signal energy measured by amplitude variation; signal frequency measured by mean zero-crossings) and nonlinear (signal complexity measured by the pattern match regularity statistic and the short-term maximum Lyapunov exponent) brain EEG dynamics in 4- and 20-month-old F344 rats with and without brain infarction. The main findings of the study were: 1) dynamic resetting of both linear and nonlinear EEG characteristics occurred following SWDs; 2) animal age significantly affected the degree of dynamic resetting in all four qEEG measures: SWDs in older rats exhibited a lower degree of dynamic resetting; 3) infarction significantly affected the degree of dynamic resetting only in terms of EEG signal complexity: SWDs in infarcted rats exhibited a lower degree of dynamic resetting; and 4) in all four qEEG measures, there was no significant interaction effect between age and infarction on dynamic resetting. We conclude that recovery of the brain to its interictal state following SWDs was better in young adult animals compared with aged animals, and to a lesser degree, in age-matched controls compared with infarction-injured animal groups, suggesting possible effects of brain resetting mechanisms and/or the disruption of the epileptogenic network that triggers SWDs., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Objective: Paraoxonase (PON) is an antioxidant enzyme linked with cardiovascular disease (CVD), diabetes as it prevents LDL oxidation. The relation of PON with the other established risk factor of diabetic complications has not been looked into., Research Design and Methods: 370 subjects were included in the study. Dividing into four group, i.e. group I included type II DM (n=220), group II was age matched control (n=100), group III were type I DM (n=25) and group IV (n=25) were age matched control group. The protocol of the study was approved by the ethical committee of the institute. SOD, GSH, PON (paraoxonase and arylesterase activity), GHb, and MDA were estimated., Results: A highly significant decrease in paraoxonase and arylesterase activity was seen in the type II DM (p<0.0001) while in type I DM both the activity was not significant (p>0.05). Paraoxonase and arylesterase activity of PONI showed a negative significant correlated with MDA (r=-0.51, p<0.0001 and r=-0.23, p<0.001) in type II DM but was not correlated in type I DM. The GHb and MDA levels were significantly increased (p<0.0001) while the levels of SOD and GSH have been decreased in type I and type II DM., Conclusion: PONI is definitely associated with development of the complications of diabetes. This may be due to the role of it as an antioxidant. As it also show a negative correlation with MDA like the other antioxidants studied., (Copyright © 2012 Diabetes India. Published by Elsevier Ltd. All rights reserved.)

Aim: To analyze the haplotype of the Ezhava population of Kerala, south India, using 8 short tandem repeat (STR) loci on the Y chromosome and trace the paternal genetic lineage of the population., Methods: Whole blood samples (n=104) were collected from unrelated healthy men of the Ezhava population over a period of one year from October 2009. Genomic DNA was extracted by salting out method. All samples were genotyped for the 8 Y-STR loci by the AmpFiSTR Y-filer PCR Amplification Kit. The haplotype and allele frequencies were determined by direct counting and analyzed using Arlequin 3.1 software, and molecular variance was calculated with the Y-chromosome haplotype reference database online analysis tool, www.yhrd.org., Results: Among the 104 examined haplotypes, we found 98 unique ones. The average gene diversity was 0.669, with the highest diversity of 0.9462 observed for the biallelic Y-STR marker DYS 385. The allele frequency among DYS loci varied between 0.0096 and 0.75. Out of the 104 haplotypes, 10 were identical to the Jat Sikh population of Punjab, which is the greatest number among the Indian populations, and 4 to the Turkish population, which is the greatest number among the European populations. According to the allele frequency of Y-STR, the Ezhavas were genetically more similar to the Europeans (60%) than to the East Asians (40%)., Conclusion: The vast majority of haplotypes were observed only once, reflecting the enormous genetic heterogeneity of the Ezhavas. Based on the genotype, the Ezhavas showed more resemblance to Jat Sikh population of Punjab and the Turkish populations than to the East Asians, hence indicating a paternal lineage of European origin.

New elastographic techniques such as poroelastography and viscoelasticity imaging aim at imaging the temporal mechanical behavior of tissues. These techniques usually involve the use of curve fitting methods being applied to noisy data to estimate new elastographic parameters. As of today, however, current elastographic implementations of poroelastography and viscoelasticity imaging methods are in general too slow and not optimized for clinical applications. Furthermore, image quality performance of these new elastographic techniques is still largely unknown due to a paucity of data and the lack of systematic studies that analyze their performance limitations. In this paper, we propose a new elastographic time constant (TC) estimator, which is based on the use of the least square error (LSE) curve-fitting method and the Levenberg-Marquardt (LM) optimization rule as applied to noisy elastographic data obtained from a material in a creep-type experiment. The algorithm is executed on a massively parallel general purpose graphics processing unit (GPGPU) to achieve real-time performance. The estimator's performance is analyzed using simulations. Experimental results obtained from poroelastic phantoms are presented as a proof of principle of the new estimator's technical applicability on real experimental data. The results of this study demonstrate that the newly proposed elastographic estimator can produce highly accurate and sensitive elastographic TC estimates in real-time and at high signal-to-noise ratios.

Staphylococci, in particular Staphylococcus aureus, are the predominant cause of bone infections worldwide. These infections are painful, debilitating and with the rise in antibiotic-resistant forms, increasingly difficult to treat. The growth in the number of prosthetic joint replacement procedures also provides new opportunities for these infections to take hold. Comprehending the mechanisms by which staphylococci interact with and damage bone is critical to the development of new approaches to meet this challenge. This review summarises current understanding of the mechanisms by which staphylococci infect and damage bone. We address the role of the inflammatory response to staphylococcal infection in disrupting the homeostatic balance of bone matrix deposition and resorption and thereby mediating bone destruction. A number of virulence factors that have been shown to contribute to bone infection and pathology are discussed, however no single factor has been defined as being specific to bone infections. Although traditionally considered an extracellular pathogen, there is increasing evidence that staphylococci are able to invade host cells, and that an intracellular lifestyle may facilitate long-term persistence in bone tissue, enabling evasion of antimicrobials and host immune responses. 'Small colony variant' strains, with mutations disabling the electron transport pathway appear particularly adept at invading and persisting within host cells, and exhibit enhanced antimicrobial resistance, and may represent a further complication in the treatment and management of staphylococcal bone disease., (Copyright 2009 Elsevier GmbH. All rights reserved.)

Background: One of the limitations of antibiotic therapy is that even after successful killing of the infecting microorganism, virulence factors may still be present and cause significant damage to the host. Light-activated antimicrobials show potential for the treatment of topical infections; therefore if these agents can also inactivate microbial virulence factors, this would represent an advantage over conventional antibiotic therapy. Staphylococcus aureus produces a wide range of virulence factors that contribute to its success as a pathogen by facilitating colonisation and destruction of host tissues., Results: In this study, the ability of the light-activated antimicrobial agent methylene blue in combination with laser light of 665 nm to inactivate staphylococcal virulence factors was assessed. A number of proteinaceous virulence factors were exposed to laser light in the presence of methylene blue and their biological activities re-determined. The activities of V8 protease, alpha-haemolysin and sphingomyelinase were shown to be inhibited in a dose-dependent manner by exposure to laser light in the presence of methylene blue., Conclusion: These results suggest that photodynamic therapy could reduce the harmful impact of preformed virulence factors on the host.

Objectives: It has been demonstrated that the efficiency of lethal photosensitization can be improved by covalently binding photosensitizing agents to bacteriophage. In this study we have investigated whether a bacteriophage requires the capacity to infect the bacterium to enhance lethal photosensitization when linked to a photosensitizer., Methods: Tin (IV) chlorin e6 (SnCe6) was conjugated to bacteriophage Phi11, a transducing phage that can infect Staphylococcus aureus NCTC 8325-4, but not epidemic methicillin-resistant S. aureus (EMRSA)-16. The conjugate and appropriate controls were incubated with these bacteria and either exposed to laser light at 632.8 nm or kept in the dark., Results: The SnCe6/Phi11 conjugate achieved a statistically significant reduction in the number of viable bacteria of both 8325-4 and EMRSA-16 strains by 2.31 log(10) and 2.63 log(10), respectively. The conjugate could not however instigate lethal photosensitization of Escherichia coli. None of the other combinations of controls, such as an equivalent concentration of SnCe6 only, an equivalent titre of bacteriophage only or experiments conducted without laser light, yielded significant reductions in the number of viable bacteria recovered., Conclusions: The inability of a bacteriophage to infect S. aureus does not prevent it from specifically delivering a photosensitizer to a bacterium enabling its lethal photosensitization.

Analysis of intracranial electroencephalographic (iEEG) recordings in patients with temporal lobe epilepsy (TLE) has revealed characteristic dynamical features that distinguish the interictal, ictal, and postictal states and inter-state transitions. Experimental investigations into the mechanisms underlying these observations require the use of an animal model. A rat TLE model was used to test for differences in iEEG dynamics between well-defined states and to test specific hypotheses: 1) the short-term maximum Lyapunov exponent (STL(max)), a measure of signal order, is lowest and closest in value among cortical sites during the ictal state, and highest and most divergent during the postictal state; 2) STL(max) values estimated from the stimulated hippocampus are the lowest among all cortical sites; and 3) the transition from the interictal to ictal state is associated with a convergence in STL(max) values among cortical sites. iEEGs were recorded from bilateral frontal cortices and hippocampi. STL(max) and T-index (a measure of convergence/divergence of STL(max) between recorded brain areas) were compared among the four different periods. Statistical tests (ANOVA and multiple comparisons) revealed that ictal STL(max) was lower (p<0.05) than other periods, STL(max) values corresponding to the stimulated hippocampus were lower than those estimated from other cortical regions, and T-index values were highest during the postictal period and lowest during the ictal period. Also, the T-index values corresponding to the preictal period were lower than those during the interictal period (p<0.05). These results indicate that a rat TLE model demonstrates several important dynamical signal characteristics similar to those found in human TLE and support future use of the model to study epileptic state transitions.

Background: The widespread problem of antibiotic resistance in pathogens such as Staphylococcus aureus has prompted the search for new antimicrobial approaches. In this study we report for the first time the use of a light-activated antimicrobial agent, methylene blue, to kill an epidemic methicillin-resistant Staphylococcus aureus (EMRSA-16) strain in two mouse wound models., Results: Following irradiation of wounds with 360 J/cm(2) of laser light (670 nm) in the presence of 100 microg/ml of methylene blue, a 25-fold reduction in the number of viable EMRSA was seen. This was independent of the increase in temperature of the wounds associated with the treatment. Histological examination of the wounds revealed no difference between the photodynamic therapy (PDT)-treated wounds and the untreated wounds, all of which showed the same degree of inflammatory infiltration at 24 hours., Conclusion: The results of this study demonstrate that PDT is effective at reducing the total number of viable EMRSA in a wound. This approach has promise as a means of treating wound infections caused by antibiotic-resistant microbes as well as for the elimination of such organisms from carriage sites.

Animal Models are used extensively in basic epilepsy research. In many studies, there is a need to accurately score and quantify all epileptic spike and wave discharges (SWDs) as captured by electroencephalographic (EEG) recordings. Manual scoring of long term EEG recordings is a time-consuming and tedious task that requires inordinate amount of time of laboratory personnel and an experienced electroencephalographer. In this paper, we adapt a SWD detection algorithm, originally proposed by the authors for absence (petit mal) seizure detection in humans, to detect SWDs appearing in EEG recordings of Fischer 334 rats. The algorithm is robust with respect to the threshold parameters. Results are compared to manual scoring and the effect of different threshold parameters is discussed.

Because of the increasing resistance of bacteria to antibiotics there is considerable interest in light-activated antimicrobial agents (LAAAs) as alternatives to antibiotics for treating localized infections. The purpose of this study was to determine whether CdSe/ZnS quantum dots (QD) could enhance the antibacterial activity of the LAAA, toluidine blue O (TBO). Suspensions of Staphylococcus aureus and Streptococcus pyogenes were exposed to white light (3600 lux) and TBO (absorbance maximum = 630 nm) in the presence and absence of 25 nm diameter QD (emission maximum = 627 nm). When the TBO:QD ratio was 2667:1, killing of Staph. aureus was enhanced by 1.72log(10) units. In the case of Strep. pyogenes, an enhanced kill of 1.55log(10) units was achieved using TBO and QD in the ratio 267:1. Singlet oxygen and fluorescence measurements showed that QD suppress the formation of singlet oxygen from TBO and that QD fluorescence is significantly quenched in the presence of TBO (70-90%). Enhanced killing appears to be attributable to a non-Förster resonance energy transfer mechanism, whereby the QD converts part of the incident light to the absorption maximum for TBO; hence more light energy is harvested, resulting in increased concentrations of bactericidal radicals. QD may, therefore, be useful in improving the efficacy of antimicrobial photodynamic therapy.

Electrochemical sensing techniques are increasingly used to study biological processes by monitoring concentration changes of the molecule of interest close to cells. The measured concentration is the result of cellular transport across the cell membrane and diffusion of the released molecules from the cells to the sensing electrode. The objective of such experiments is to understand the cellular processes underlying the observed changes in concentration. Thus, the influence of mass transport on the measured concentration trace has to be removed. This is done by deconvolution of the impulse response function of diffusion from the concentration data. We have recently observed that measuring concentration at a sampling rate that satisfies the Nyquist criterion for the observed concentration dynamics may not be sufficient to correctly reconstruct cellular flux. This is because the impulse response function of diffusion also has to be represented with sufficient temporal resolution. We discuss this problem here using the example of monitoring drug efflux from a monolayer of cancer cells with microvoltammetry, and chloride secretion from an epithelial cell monolayer monitored with an ion-selective electrode.

To understand the increase in age-related incidence and frequency of absence seizures in the rat brain, we investigated the effect of these seizures on brain dynamics. This paper puts forward the hypothesis that age-related differences in the expression of absence seizures are associated with the ability of the seizures to reset brain dynamics.

There has been much recent interest in alpha-1,3-glucanases (mutanases) as they have the potential to be used in the treatment of dental caries. Mutanases have been reported in a number of bacteria, yeast and fungi but remain a relatively uncharacterised family of enzymes. In this study we heterologously expressed the mutanase gene from the filamentous fungus Penicillium purpurogenum to enable further characterization of its enzymatic activity. The mutanase cDNA was cloned and expressed in the methylotrophic yeast Pichia pastoris. The molecular mass of the secreted protein was about 102 kDa. The recombinant enzyme hydrolyzed mutan with a specific activity of 3.9 U/mg of protein. The recombinant enzyme was specific for mutan and could not cleave a variety of other polysaccharides demonstrating a specificity for alpha-1,3-glucosidic linkages. The pH and temperature optima were pH 4.6 and 45 degrees C, respectively. Synthetic compounds were also tested as substrates to assess whether the P. purpurogenum mutanase has an exo- or endo-type mechanism of hydrolysis. The results suggest an endo-hydrolytic mode of action. The type of mechanism was confirmed since mutanase activity was not suppressed in the presence of inhibitors of exo-type enzymes.

Background: The increase in resistance to antibiotics among disease-causing bacteria necessitates the development of alternative antimicrobial approaches such as the use of light-activated antimicrobial agents (LAAAs). Light of an appropriate wavelength activates the LAAA to produce cytotoxic species which can then cause bacterial cell death via loss of membrane integrity, lipid peroxidation, the inactivation of essential enzymes, and/or exertion of mutagenic effects due to DNA modification. In this study, the effect of the LAAA indocyanine green excited with high or low intensity light (808 nm) from a near-infrared laser (NIR) on the viability of Staphylococcus aureus, Streptococcus pyogenes and Pseudomonas aeruginosa was investigated., Results: All species were susceptible to killing by the LAAA, the bactericidal effect being dependent on both the concentration of indocyanine green and the light dose. Indocyanine green photosensitization using both high (1.37 W cm(-2)) and low (0.048 W cm(-2)) intensity NIR laser light was able to achieve reductions of 5.6 log10 (>99.99%) and 6.8 log10 (>99.99%) in the viable counts of Staph. aureus and Strep. pyogenes (using starting concentrations of 106-107 CFU ml(-1)). Kills of 99.99% were obtained for P. aeruginosa (initial concentration 108-109 CFU ml(-1)) photosensitized by the high intensity light (1.37 W cm(-2)); while a kill of 80% was achieved using low intensity irradiation (0.07 W cm(-2)). The effects of L-tryptophan (a singlet oxygen scavenger) and deuterium oxide (as an enhancer of the life span of singlet oxygen) on the survival of Staph. aureus was also studied. L-tryptophan reduced the proportion of Staph. aureus killed; whereas deuterium oxide increased the proportion killed suggesting that singlet oxygen was involved in the killing of the bacteria., Conclusion: These findings imply that indocyanine green in combination with light from a near-infrared laser may be an effective means of eradicating bacteria from wounds and burns.

Successful systemic management of stage IV melanoma continues to be elusive because of the paucity of effective therapies. This has fueled the continued interest in surgical resection. Several single-institution studies and a current, large, multi-institutional phase III trial have demonstrated a survival benefit for patients who underwent surgical resection for melanoma metastases. Incorporating these results into new approaches using multimodality treatment may enhance survival in patients with stage IV melanoma., ((Copyright) 2008 Wiley-Liss, Inc.)

Phage display screening with fragmented genomic DNA from the animal pathogen Pasteurella multocida has identified a gene encoding a putative fibronectin binding protein (19). Homologues of this gene (PM1665) are found in all other sequenced members of the Pasteurellaceae. Gene PM1665 has been cloned, and the protein has been expressed. Recombinant PM1665 protein binds to both soluble and immobilized fibronectin and is unique in that it interacts with the integrin-binding fibronectin type III (FnIII) repeats FnIII(9-10) and not, as is the case for almost all other fibronectin adhesins, to the N-terminal type I repeats. Surface plasmon resonance analysis revealed a complex binding mechanism with a K(D) (equilibrium dissociation constant) of 150 nM +/- 70 nM. Bioinformatics analysis suggests that the PM1665 protein contains two helix-hairpin-helix (HhH) motifs, and truncation mutation studies have identified the binding site in the protein as a combination of these two HhH motifs in conjunction with a conserved amino acid motif, VNINTA. We have shown that the PM1665 protein is on the cell surface and that binding of P. multocida to fibronectin is almost completely inhibited by anti-PM1665 antiserum. These results support the hypothesis that the PM1665 protein is a member of a new family of fibronectin binding adhesins that are important in the adhesion of P. multocida to fibronectin.

A novel fibronectin-binding protein from Pasteurella multocida (PM1665) that binds to the fibronectin type III(9-10) modules via two helix-hairpin-helix motifs has recently been described [1]. This protein shares homology with competence-related DNA-binding and uptake proteins (ComEA and ComE) from Gram-positive and Gram-negative bacteria. Here, we show that recombinant PM1665 (now designated ComE1) also binds to DNA through the same helix-hairpin-helix motifs required for fibronectin-binding. This binding to DNA is non sequence-specific and is confined to double-stranded DNA. We have cloned and expressed ComE1 proteins from five members of the Pasteurellaceae in order to further investigate the function(s) of these proteins. When expressed as recombinant GST-fusion proteins, all of the homologues bound both to fibronectin and to double-stranded DNA. Inactivation of the gene encoding the ComE1 homologue in Actinobacillus pleuropneumoniae indicates major roles for these proteins in at least two processes: natural transformation, and binding of bacteria to fibronectin.

The Pasteurellaceae contain a number of important animal pathogens. Although related, the various members of this family cause a diversity of pathology in a wide variety of organ systems. Adhesion is an important virulence factor in bacterial infections. Surprisingly little is known about the adhesins of the Pasteurellaceae. To attempt to identify the genes coding for adhesins to some key components of the hosts extracellular matrix molecules, phage display libraries of fragmented genomic DNA from Haemophilus influenzae, Actinobacillus pleuropneumoniae, Pasteurella multocida and Aggregatibacter actinomycetemcomitans, were prepared in the phage display vector pG8SAET. The libraries were screened against human or porcine fibronectin, serum albumin or a commercial extracellular matrix containing type IV collagen, laminin and heparin sulphate. Four genes encoding putative adhesins were identified. These genes code for: (i) a 34 kDa human serum albumin binding protein from Haemophilus influenzae; (ii) a 12.8 kDa fibronectin-binding protein from Pasteurella multocida; (iii) a 13.7 kDa fibronectin-binding protein from A. actinomycetemcomitans; (iv) a 9.5 kDa serum albumin-binding protein from A. pleuropneumoniae. None of these genes have previously been proposed to code for adhesins. The applications of phage display with whole bacterial genomes to identify genes encoding novel adhesins in this family of bacteria are discussed.

Background: Circulating fetal cells and cell free DNA in the maternal blood has been shown to help in prenatal diagnosis of genetic disorders without relying on invasive procedures leading to significant risk of pregnancy loss., Aim: The current study was undertaken to detect the male fetal population using Y STR markers DYS 19, DYS 385 and DYS 392 and also to study the extent of persistence of fetal DNA in the mother following delivery., Materials and Methods: Blinded study was conducted on 50 mothers delivering male and female babies. Cellular and cell free DNA was extracted from maternal and fetal cord blood and amplified for Y STR markers by PCR., Results: The amplification sensitivity of Y specific STR, DYS19 was 100% (22/22) in the male fetal DNA samples. The incidence of other STRs, i.e., DYS385 and DYS392 were 91% (20/22) each. Analysis of results revealed that thirteen of the twenty six women had detectable male fetal DNA at the time of delivery. However fetal DNA was not detectable twenty four hours after delivery., Conclusion: Preliminary results show that the separation of fetal cell-free DNA in the maternal circulation is a good low-cost approach for the future development of novel strategies to provide non-invasive techniques for early prenatal diagnosis.

Bone implants infected with Staphylococcus epidermidis often require surgical intervention because of the failure of antibiotic treatment. The reasons why such infections are resistant to therapy are poorly understood. We have previously reported that another bacterium, Staphylococcus aureus, can invade bone cells and thereby evade antimicrobial therapy. In this study we have investigated the hypothesis that S. epidermidis can also invade bone cells and may therefore explain the difficulties of treating infections with this organism. We found that S. epidermidis was capable of invading bone cells but that there were significant strain dependent differences in this capacity. A recombinant protein encompassing the D1-D4 repeat region of S. aureus fibronectin-binding protein B completely inhibited internalization of S. aureus but failed to block internalization of S. epidermidis. Similarly a blocking antibody to alpha5beta1 integrin inhibited internalization of S. aureus by bone cells but had no effect on the uptake of S. epidermidis. Therefore unlike S. aureus, S. epidermidis does not gain entrance into bone cells through a fibronectin bridge between the alpha5beta1 integrin and a bacterial adhesin.

A 39-year-old man presented with a chronic nonhealing toe ulcer and multiple skin-colored papules on the back, arms, and knees. From an initial small erosion, the toe lesion ulcerated over a 1.5-year period, while the papules progressed over a 6-month period, first appearing on the back and then spreading to the arms and knees. The past medical and family history were non-contributory. Pertinent findings included the aforementioned well-defined, asymmetric, shiny papules (Fig. 1). There were multiple, discrete, erythematous, pedunculated, tumor-like masses of various sizes over the right thigh (Fig. 2) and a well-defined 4 cm x 3 cm ulcer on the second toe of the left foot, the floor of which was covered by necrotic slough. There was distal loss of sensation to temperature, touch, and pain. The greater auricular, ulnar, radial cutaneous and common peroneal nerves were thickened bilaterally, but non-tender. A clinical diagnosis of histoid leprosy was made. The differential diagnosis for the tumor-like thigh masses included dermatofibroma, neurofibroma, and Kaposi's sarcoma. The hemogram, liver/renal function tests, chest X-ray, and abdominal ultrasound were normal. Human immunodeficiency virus enzyme-linked immunosorbent assay (HIV ELISA) was negative. The ear lobe smear (ELS) for acid-fast bacilli showed a bacterial index (BI) of 6+[> 1000 organisms/oil immersion field (oif)] and a morphological index (MI) of 50%. The skin-colored papules on the back and the pedunculated masses showed a BI of 5+ (100-1,000 organisms/oif) and an MI of 50%. The normal skin showed a BI of 4+ (10-100 organisms/oif) and an MI of 5%. Biopsy of the papules and pedunculated tumors showed a well-circumscribed area of the dermis packed with many acid-fast organisms and foamy macrophages, consistent with histoid leprosy (Fig. 3). Fite-Faraco stain demonstrated cells packed with lepra bacilli. A final diagnosis of lepromatous leprosy, histoid variant, was made. Dapsone, clofazimine, rifampicin, and, later, ofloxacin were started.

Microbial infections are dependent on the panoply of interactions between pathogen and host and identifying the molecular basis of such interactions is necessary to understand and control infection. Phage display is a simple functional genomic methodology for screening and identifying protein-ligand interactions and is widely used in epitope mapping, antibody engineering and screening for receptor agonists or antagonists. Phage display is also used widely in various forms, including the use of fragment libraries of whole microbial genomes, to identify peptide-ligand and protein-ligand interactions that are of importance in infection. In particular, this technique has proved successful in identifying microbial adhesins that are vital for colonization.

Progressive preictal dynamical convergence and postictal divergence of dynamical EEG descriptors among brain regions has been reported in human temporal lobe epilepsy (TLE) and in a rodent model of TLE. There are also reports of anticonvulsant effects of high frequency stimulation of the hippocampus in humans. We postulate that this anticonvulsant effect is due to dynamical resetting by the electrical stimulation. The following study investigated the effects of acute hippocampal electrical stimulation on dynamical transitions in the brain of a spontaneously seizing animal model of TLE to test the hypothesis of divergence in dynamical values by electrical stimulation of the hippocampus.

Light-activated antimicrobial agents (photosensitizers) are promising alternatives to antibiotics for the treatment of topical infections. To improve efficacy and avoid possible damage to host tissues, targeting of the photosensitizer to the infecting organism is desirable, and this has previously been achieved using antibodies and chemical modification of the agent. In this study we investigated the possibility of using a bacteriophage to deliver the photosensitizer tin(IV) chlorin e6 (SnCe6) to Staphylococcus aureus. SnCe6 was covalently linked to S. aureus bacteriophage 75, and the ability of the conjugate to kill various strains of S. aureus when exposed to red light was determined. Substantial kills of methicillin- and vancomycin-intermediate strains of S. aureus were achieved using low concentrations of the conjugate (containing 1.5 microg/ml SnCe6) and low light doses (21 J/cm2). Under these conditions, the viability of human epithelial cells (in the absence of bacteria) was largely unaffected. On a molar equivalent basis, the conjugate was a more effective bactericide than the unconjugated SnCe6, and killing was not growth phase dependent. The conjugate was effective against vancomycin-intermediate strains of S. aureus even after growth in vancomycin. The results of this study have demonstrated that a bacteriophage can be used to deliver a photosensitizer to a target organism, resulting in enhanced and selective killing of the organism. Such attributes are desirable in an agent to be used in the photodynamic therapy of infectious diseases.

Staphylococcus aureus produce a family of exotoxins (staphylococcal superantigen like proteins, SSLs) with structural, but not functional, homology to superantigens. These proteins have previously been shown to interact selectively with antigen presenting cells, including dendritic cells. The functional consequences of this interaction are now explored. SSL7 and 9 had no effect on viability or morphology of dendritic cells. The proteins did not induce dendritic cell maturation, as measured by cell surface phenotype. Exposure to SSL did not alter the ability of dendritic cells to take up FITC-dextran. Finally, exposure to SSLs did not impair the ability of the dendritic cells to stimulate allogeneic or antigen specific T cell responses. However, dendritic cells loaded with SSL7 or 9 were able to stimulate a T cell proliferative response in 3/8 healthy individuals tested. Sera from nine out of 10 individuals tested contained antibodies against both SSL7 and SSL9, and the response to each SSL was specific and not cross-reactive. The results demonstrate that SSLs are immunogenic in humans at both the B and T cell level, but it remains unclear whether this response is to the benefit of the bacterium or the host.

Vaccination of the susceptible livestock with potent, safe and cost effective vaccine is the primary requirement to control foot-and-mouth disease (FMD) in an endemic country. In this study, an alternative approach was used in which structural protein genes of all the four serotypes of FMDV (O, Asia 1, A22 and C) were expressed separately in methylotrophic yeast Pichia pastoris. The recombinant polyproteins (P1) were characterized by SDS-PAGE and in Western Blot analysis. Partially purified protein was used for immunization in guinea pigs with different adjuvant formulations and immune response studied. Ninety micrograms of the recombinant protein per monovalent dose was used for immunization. A single injection of a monovalent or polyvalent vaccine was given to guinea pigs with various adjuvant combinations viz., Monovalent recombinant protein either adjuvanted with Montanide-ISA50V or Indigenous oil, Monovalent recombinant protein mixed with 1/10th dose of inactivated oil-adjuvanted virus vaccine and Polyvalent recombinant protein with Montanide ISA50V. FMDV specific humoral immune response was observed at about 28th day post vaccination. The immune response as assessed by indirect ELISA and Serum neutralization test titres was found to be 320-640 and 16-32, respectively. When challenged with virulent homologous type 'O' virus, the guinea pigs showed protective C index of 2.01,1.81, 2.56 and 2.48, respectively, with above said adjuvant combinations. The study has shown that yeast-expressed FMDV P1 polyprotein in a single dose could elicit a protective immune response in guinea pigs, and this could be a possible future vaccine candidate in homologous host.

Cytolethal distending toxin (CDT), which is encoded by three genes, cdtA, cdtB and cdtC, is now recognized to have a growing list of biological actions, including inhibition of cell cycle progression, promotion of apoptosis and stimulation of cytokine secretion. It appears that internalization of CDT is essential, at least for cell cycle blockade. Using purified recombinant CDT proteins from the periodontopathic bacterium Actinobacillus actinomycetemcomitans, the authors investigated which combination of toxin proteins produce cell cycle inhibition and which bound and/or entered into host cells. No evidence was found that CdtB bound to HEp-2 human epithelial cells. In contrast, both CdtA and CdtC bound to these cells. Induction of cell cycle arrest required that cells be exposed to both CdtB and CdtC. Pre-exposure of cells to CdtC for as little as 10 min, followed by removal of the free CdtC and addition of exogenous CdtB, resulted in the inhibition of cell cycle progression, suggesting that CdtB could bind to cell-surface-located CdtC. Using various methods to follow internalization of the CDT proteins it was concluded that CdtC acts to bind CdtB at the cell surface and transports it into the cell as a complex via an endosomal pathway blockable by monensin and brefeldin A.

Cytolethal distending toxin (CDT) is a novel bacterial toxin that is produced by a variety of pathogenic bacteria. The mechanism of cytotoxicity of CDT is unique in that it enters into eukaryotic cells and breaks double-stranded DNA. This initiates the cell's own DNA damage-response mechanisms, resulting in the arrest of the cell cycle at the G2/M boundary. Affected cells enlarge until they finally undergo programmed cell death. This review encompasses recent work on CDT and focuses on the molecular mechanisms used by this toxin to block cell-cycle progression, the benefit to the bacterium of possession of this toxin and the clinical relevance of intoxication.

We describe the results of an inelastic neutron scattering measurement of the magnetic excitations in SrCaGaMnO(5+δ), a quasi-two-dimensional compound whose structure consists of layers of MnO(6) octahedra separated by layers of GaO(4) tetrahedra (the brownmillerite structure), and Ca(2.5)Sr(0.5)Mn(2)GaO(8), a bilayered brownmillerite. In both materials, a band of magnetic scattering appears below the magnetic ordering temperature which can be associated with magnon excitations. Our measurements allow us to provide an estimate for the intraplane exchange constant in both materials, which we find to be 3.4(4) meV for SrCaGaMnO(5+δ) and 2.2(4) meV for Ca(2.5)Sr(0.5)Mn(2)GaO(8).

Staphylococcus aureus is a common wound-infecting organism which can interact with cells via the extracellular matrix protein fibronectin (FN). The aim of this study was to determine the effect of the FN-binding protein (FnBP) of S. aureus on the behaviour of a human skin keratinocytes cell line (UP). FN-coated plates and Transwell membranes were exposed to a recombinant protein encompassing the D1-D4 repeat region of S. aureus FnBPB (rFnBPBD1-D4) before cell adhesion and migration assays. The influence of this protein on migration into a wounded area, cell cycle progression and endogenous cellular FN assembly was also assessed. The rFnBPBD1-D4 protein potently inhibited S. aureus entry into UP keratinocytes but had no effect on cell adhesion to FN substrate. It inhibited UP keratinocyte Transwell migration by 17% (P= 0.04) and 31% (P=0.02) at 10 and 100 microg/ml, respectively. In the wound assay, 100 microg/ml of rFnBPBD1-D4 protein reduced the migration area by approximately 30%. No differences in cell cycle progression were observed. In the presence of rFnBPBD1-D4, most of the cellular FN matrix on the cell surface and along the cell filopodia seen in untreated cells was absent. Interaction of S. aureus FnBPs with FN may influence cell behaviour and thus play a role in delayed epithelial closure in infected healing wounds.

The temperature dependence of the crystal structure and electronic properties of brownmillerite-like Ca(2.5)Sr(0.5)GaMn(2)O(8) has been studied by neutron powder diffraction and muSR spectroscopy. The results show that short-range 2D magnetic order begins to develop within the perovskite-like bilayers of MnO(6) octahedra approximately 50 K above the 3D Néel temperature of approximately 150 K. The bilayers show a structural response to the onset of magnetism throughout this temperature range whereas the GaO(4) layers that separate the bilayers only respond below the 3D ordering temperature. XANES spectroscopy shows that the sample contains Mn(3+) and Mn(4+) cations in a 1:1 ratio, and the behavior in the region of the Néel transition is interpreted as a local charge ordering. Electron diffraction and high-resolution electron microscopy have been used to show that the local microstructure is more complex than the average structure revealed by neutron diffraction, and that microdomains exist in which the GaO(4) tetrahedra show different orientations. It is argued that the bonding requirements of diamagnetic gallium control the electronic behavior within the perovskite-like bilayers.

Staphylococcus aureus is among the most important human pathogens and causes various superficial and systemic infections. The ability of S. aureus to be internalized by, and survive within, host cells, such as keratinocytes, may contribute to the development of persistent or chronic infections and may finally lead to deeper tissue infections or dissemination. To examine the mechanisms of internalization of S. aureus by keratinocytes, isogenic mutants lacking fibronectin-binding proteins (FnBPs), a recombinant protein consisting of the fibronectin-binding domain of S. aureus FnBPs, and an anti-alpha5beta1 antibody were used in cocultures with immortalized keratinocytes and primary keratinocytes. We found that internalization of S. aureus by immortalized keratinocytes requires bacterial FnBPs and is mediated by the major fibronectin-binding integrin alpha5beta1. In contrast to internalization by immortalized keratinocytes, internalization of S. aureus by primary keratinocytes could occur through FnBP-dependent and -independent pathways. S. aureus clumping factor B (ClfB), which was recently determined to bind to epithelial cells, was not involved in the uptake of this bacterium by keratinocytes. The identification of an alternate uptake pathway, which is independent of S. aureus FnBPs and host cell alpha5beta1, has important implications for the design of therapies targeted to bacterial uptake by host cells.

The staphylococcal superantigen-like proteins (SSLs) are a family of polymorphic paralogs encoded in the Staphylococcus aureus genome whose function is unknown. The crystal structure of SSL7 was determined and compared to that of SSL5 and that of a classical superantigen, streptococcal pyrogenic exotoxin. Although the overall architecture of the superantigen family is retained in both SSL7 and SSL5, there are significant differences in the structures which suggest that the characteristic major histocompatibility complex binding site of superantigens has been lost. To complement these data, the abilities of SSL7 and a closely related paralog, SSL9, to interact with cells of the immune system were investigated. In populations of human white blood cells, both SSLs interacted selectively with monocytes via specific saturable but separate binding sites, which led to rapid uptake of the SSLs. In addition, SSLs were rapidly taken up by dendritic cells, but not by macrophages, into the same endosomal compartment as dextran. The ability of these secreted proteins to target antigen-presenting cells may enhance a misplaced antibody response against the proteins, which may facilitate bacterial colonization rather than contribute to host protection. Like classical superantigens, therefore, SSLs may distract the host's immune system, but they may do so via entirely different molecular mechanisms.

The International Nomenclature Committee for Staphylococcal Superantigens proposes an international procedure for the designation of newly described superantigens and putative superantigens, a procedure that will be compatible with the new age of genomics.