Photocleavable Visible Light-Triggered Anthraquinone-Derived Water-Soluble Block Copolymer for Peroxynitrite Generation in Cancer Therapy.

We report a facile stimuli-responsive strategy to generate reactive oxygen and nitrogen species (ROS and RNS) in the biological milieu from a photocleavable water-soluble block copolymer under visible light irradiation (427 nm, 2.25 mW/cm2). An anthraquinone-based water-soluble polymeric nitric oxide (NO) donor (BCPx-NO) is synthesized, which exhibits NO release in the range of 40-65 µM within 10 h of photoirradiation with a half-life of 30-103 min. Additionally, BCPx-NO produces peroxynitrite (ONOO-) and singlet oxygen (1O2) under photoirradiation. To understand the mechanism of NO release and photolysis of the functional group under blue light, we prepared a small-molecule anthraquinone-based N-nitrosamine (NOD). The cellular investigation of the effect of spatiotemporally controlled ONOO- and 1O2 generation from the NO donor polymeric nanoparticles in a triple negative breast adenocarcinoma (MDA-MB-231) under visible light irradiation (white light, 5.83 mW/cm2; total dose 31.5 J/cm2) showed an IC50 of 0.6 mg/mL. The stimuli-responsive strategy using a photolabile water-soluble block copolymer employed to generate ROS and RNS in a biological setting widens the horizon for their potential in cancer therapy.

Photostimulated Extended Nitric Oxide (NO) Release from Water-Soluble Block Copolymer to Enhance Antibacterial Activity.

N-Nitrosamines are well established motifs to release nitric oxide (NO) under photoirradiation. Herein, a series of amphiphilic N-nitrosamine-based block copolymers (BCPx-NO) are developed to attain controlled NO release under photoirradiation (365 nm, 3.71 mW/cm2). The water-soluble BCPx-NO forms micellar architecture in aqueous medium and exhibits a sustained NO release of 92-160 µM within 11.5 h, which is 36.8-64.0% of the calculated value. To understand the NO release mechanism, a small molecular NO donor (NOD) resembling the NO releasing functional motif of BCPx-NO is synthesized, which displays a burst NO release in DMSO within 2.5 h. The radical nature of the released NO is confirmed by electron paramagnetic resonance (EPR) spectroscopy. The gradual NO release from micellar BCPx-NO enhances antibacterial activity over NOD and exhibits a superior bactericidal effect on Gram-positive Staphylococcus aureus. In relation to biomedical applications, this work offers a comprehensive insight into tuning light-triggered NO release to improve antibacterial activity.

Paradigm Changing Integration Technology for the Production of Flexible Electronics by Transferring Structures, Dies and Electrical Components from Rigid to Flexible Substrates.

Emerging trends like the Internet of Things require an increasing number of different sensors, actuators and electronic devices. To enable new applications, such as wearables and electronic skins, flexible sensor technologies are required. However, established technologies for the fabrication of sensors and actuators, as well as the related packaging, are based on rigid substrates, i.e., silicon wafer substrates and printed circuit boards (PCB). Moreover, most of the flexible substrates investigated until now are not compatible with the aforementioned fabrication technologies on wafers due to their lack of chemical inertness and handling issues. In this presented paper, we demonstrate a conceptually new approach to transfer structures, dies, and electronic components to a flexible substrate by lift-off. The structures to be transferred, including the related electrical contacts and packaging, are fabricated on a rigid carrier substrate, coated with the flexible substrate and finally lifted off from the carrier. The benefits of this approach are the combined advantages of using established semiconductor and microsystem fabrication technologies as well as packaging technologies, such as high precision and miniaturization, as well as a variety of available materials and processes together with those of flexible substrates, such as a geometry adaptivity, lightweight structures and low costs.

Styrene-Maleimide/Maleic Anhydride Alternating Copolymers: Recent Advances and Future Perspectives.

Alternating sequencing of styrene-maleimide/maleic anhydride (S-MI/MA) in the copolymer chain is known for a long time. But since early 2000, this class of copolymers has been extensively studied using various living/controlled polymerization techniques to design S-MI/MA alternating copolymers with tunable molecular weight, narrow dispersity (Ð), and precise chain-end functionality. The widespread diverse applications of this polymeric backbone are due to its ease of synthesis, cheap starting materials, high precision in alternating sequencing, and facile post-polymerization functionalization with simple organic reactions. Recently, S-MI/MA alternating copolymers have been rediscovered as novel polymers with unprecedented emissive behavior. It outperforms the traditional fluorophores with no aggregation caused quenching (ACQ), aqueous solubility, and greater cell viability. Herein, the origin of alternating sequence, synthesis, and recent (2010-Present) developments in applications of these polymers in different fields are elaborately discussed, including the advantages of the unconventional luminogenic property. This review article also highlights the future research directions of the versatile S-MI/MA copolymers.

Nitric Oxide Releasing Delivery Platforms: Design, Detection, Biomedical Applications, and Future Possibilities.

Gasotransmitters belong to the subfamily of endogenous gaseous signaling molecules, which find a wide range of biomedical applications. Among the various gasotransmitters, nitric oxide (NO) has an enormous effect on the cardiovascular system. Apart from this, NO showed a pivotal role in neurological, respiratory, and immunological systems. Moreover, the paradoxical concentration-dependent activities make this gaseous signaling molecule more interesting. The gaseous NO has negligible stability in physiological conditions (37 °C, pH 7.4), which restricts their potential therapeutic applications. To overcome this issue, various NO delivering carriers were reported so far. Unfortunately, most of these NO donors have low stability, short half-life, or low NO payload. Herein, we review the synthesis of NO delivering motifs, development of macromolecular NO donors, their advantages/disadvantages, and biological applications. Various NO detection analytical techniques are discussed briefly, and finally, a viewpoint about the design of polymeric NO donors with improved physicochemical characteristics is predicted.

Towards Optimal Decomposition of Boolean Networks.

In recent years, great efforts have been made to analyze biological systems to understand the long-run behaviors. As a well-established formalism for modelling real-life biological systems, Boolean networks (BNs) allow their representation and analysis using formal reasoning and tools. Most biological systems are robust-they can withstand the loss of links and cope with external environmental perturbations. Hence, the BNs used to model such systems are necessarily large and dense, and yet modular. However, existing analysis methods only work well on networks of moderate size. Thus, there is a great need for efficient methods that can handle large-scale BNs and for doing so it is inevitable to exploit both the structural and dynamic properties of the networks. In this paper, we propose a method towards the optimal decomposition of BNs to balance the relation between the structure and dynamics of a network. We show that our method can greatly improve the existing decomposition-based attractor detection by analyzing a number of large real-life biological networks.

Genome-wide unique insertion sequences among five Brucella species and demonstration of differential identification of Brucella by multiplex PCR assay.

Brucellosis is a neglected zoonotic disease caused by alpha proteobacterial genus Brucella comprising of facultative intracellular pathogenic species that can infect both animals and humans. In this study, we aimed to identify genome-wide unique insertion sequence (IS) elements among Brucella abortus, B. melitensis, B. ovis, B. suis and B. canis for use in species differentiation by conducting an intensive in silico-based comparative genomic analysis. As a result, 25, 27, 37, 86 and 3 unique ISs were identified respectively and they had a striking pattern of distribution among them. To explain, a particular IS would be present in four species with 100% identity whereas completely absent in the fifth species. However, flanking regions of that IS element would be highly identical and conserved in all five species. Species-specific primers designed on these flanking conserved regions resulted in two different amplicons grouping the species into two: one that possesses IS and the other that lacks it. Seeking for species-specific amplicon size for particular species was sufficient to identify it irrespective of biovar. A multiplex PCR developed using these primers resulted in successful differentiation of the five species irrespective of biovars with significant specificity and sensitivity when examined on clinical samples.

An Efficient Approach Towards the Source-Target Control of Boolean Networks.

We study the problem of computing a minimal subset of nodes of a given asynchronous Boolean network that need to be perturbed in a single-step to drive its dynamics from an initial state to a target steady state (or attractor), which we call the source-target control of Boolean networks. Due to the phenomenon of state-space explosion, a simple global approach that performs computations on the entire network may not scale well for large networks. We believe that efficient algorithms for such networks must exploit the structure of the networks together with their dynamics. Taking this view, we derive a decomposition-based solution to the minimal source-target control problem which can be significantly faster than the existing approaches on large networks. We then show that the solution can be further optimized if we take into account appropriate information about the source state. We apply our solutions to both real-life biological networks and randomly generated networks, demonstrating the efficiency and efficacy of our approach.

Controlling large Boolean networks with single-step perturbations.

MOTIVATION: The control of Boolean networks has traditionally focussed on strategies where the perturbations are applied to the nodes of the network for an extended period of time. In this work, we study if and how a Boolean network can be controlled by perturbing a minimal set of nodes for a single-step and letting the system evolve afterwards according to its original dynamics. More precisely, given a Boolean network (BN), we compute a minimal subset Cmin of the nodes such that BN can be driven from any initial state in an attractor to another 'desired' attractor by perturbing some or all of the nodes of Cmin for a single-step. Such kind of control is attractive for biological systems because they are less time consuming than the traditional strategies for control while also being financially more viable. However, due to the phenomenon of state-space explosion, computing such a minimal subset is computationally inefficient and an approach that deals with the entire network in one-go, does not scale well for large networks. RESULTS: We develop a 'divide-and-conquer' approach by decomposing the network into smaller partitions, computing the minimal control on the projection of the attractors to these partitions and then composing the results to obtain Cmin for the whole network. We implement our method and test it on various real-life biological networks to demonstrate its applicability and efficiency. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Algorithms for the Sequential Reprogramming of Boolean Networks.

Cellular reprogramming, a technique that opens huge opportunities in modern and regenerative medicine, heavily relies on identifying key genes to perturb. Most of the existing computational methods for controlling which attractor (steady state) the cell will reach focus on finding mutations to apply to the initial state. However, it has been shown, and is proved in this article, that waiting between perturbations so that the update dynamics of the system prepares the ground, allows for new reprogramming strategies. To identify such sequential perturbations, we consider a qualitative model of regulatory networks, and rely on Binary Decision Diagrams to model their dynamics and the putative perturbations. Our method establishes a set identification of sequential perturbations, whether permanent (mutations) or only temporary, to achieve the existential or inevitable reachability of an arbitrary state of the system. We apply an implementation for temporary perturbations on models from the literature, illustrating that we are able to derive sequential perturbations to achieve trans-differentiation.

A novel tandem repeat cloning technique for creation of multiple short peptide repeats to differentiate closely related antigens.

Antibody cross-reactivity is a problem often associated with closely related antigens. This study was aimed to develop a method enabling differentiation of closely related toxins based on antigen designing strategy. The method involves identification of disparate amino acids (AA) confined to target antigen in comparison with two or more closely related antigens, their assembly into a DNA oligomer and further cloning as six tandem repeats (TR) using restriction and ligation strategy into a desired vector and finally generation of antigen specific antibodies. The practical utility of this method was demonstrated by generating and testing the specificity of polyclonal antibodies against staphylococcal enterotoxin C (SEC). Cross-reactivity is a problem often associated with SEC in immunoassays due to its amino acid sequence identity with staphylococcal enterotoxin B (SEB) (40-60%). To circumvent the same, the above-mentioned strategy was applied. Unique AA of SEC (36 AA) in comparison to SEB were selected, reassembled and with deduced corresponding nucleotides, an oligomer of 117 bases was designed. Using primers with restriction overhangs, three constructs were created each with two repeats using a common restriction site. The resulting three constructs were sequentially cloned into alternating restriction sites of pRSET A vector in directional orientation, expressed in E. coli for rTR/SEC protein which was used to generate specific polyclonal antibodies against SEC. Specificity was compared with antibody raised against whole SEC recombinant protein using Western blot and dot blot assays. High specificity was achieved through the developed strategy signifying its possible application to address cross-reactivity problem associated with closely related antigens.

Protective and therapeutic efficacy study of divalent fusion protein rL7/L12-Omp25 against B. abortus 544 in presence of IFNγ.

Brucella as intracellular pathogen requires a coordinate interaction between Th1 subset of gamma interferon-secreting CD4 T cells and CD8 T cells for optimal protective immunity. It was previously recognized that L7/L12 as T cell-reactive antigen from the pathogen. On other hand, Omp25 was found as another antigen to provide protection against the Brucella infection by eliciting both Th1 and Th2 type of immune responses in mice. Here, we analyzed the prophylactic and therapeutic efficacy of a divalent fusion protein (rL7/L12-Omp25) comprising these two promising immunogens of Brucella in the presence of murine IFN-gamma in mice against B. abortus 544 challenge. rIFN-gamma with rL7/L12-Omp25 resulted in superior immune response when compared to the animal vaccine strain B. abortus S19. The vaccine candidate caused dominance of IgG1 over IgG2a and upregulated cytokine secretion (IFN-gamma, TNF-α, and IL-10) among immunized mice. Moreover, the antigen in combination with murine IFN-gamma elicited stronger cell-mediated immune response among the immunized animals when compared to standard vaccine (S19). The registered log protection unit among challenged mice with B. abortus 544 pathogen was 2.16, p = 0.0001 when rL7/L12-Omp25 was administered alone and 2.4, p = 0.0001 when it was administered along with rIFN-gamma. However, the molecule upon administration with murine IFN-gamma imparted very minimal or no therapeutic effect against brucellosis. To conclude, our study demonstrates the potential of rL7/L12-Omp25 as an immunogen of prospective and efficient prophylaxis as it is capable of eliciting both cell-mediated and humoral immune responses against brucellosis.

A simple end-point assay for calcium channel activity.

Cellular calcium signaling events are transient. Hence they are observed in real time using fluorescence imaging or electrophysiological methods that require sophisticated instrumentation and specialized skills. For high throughput assays simple and inexpensive techniques are desirable. Many calcium channels that serve as drug targets have subtypes arising from diverse subunit combinations. These need to be targeted selectively for achieving efficacy and for avoiding side effects in therapies. This in turn increases the number of calcium channels that act as drug targets. We report a novel method for intracellular calcium sensing that utilizes the calcium dependent stable interaction between CaM kinase II (CaMKII) and its ligands such as the NMDA receptor subunit GluN2B. The CaMKII-GluN2B complex formed persists as a memory of the transient increase in calcium. In a cell-based assay system GFP-α-CaMKII expressed in the cytosol responds to calcium by translocating towards GluN2B sequence motif exogenously expressed on mitochondria or endoplasmic reticulum. The resulting punctate fluorescence pattern serves as the signal for intracellular calcium release. The pattern is stable, unaffected by sample processing and is observable without real time imaging. The activities of calcium channel proteins heterologously expressed in HEK-293 cells were detected with specificity using this technique. A calcium sensor vector and a calcium sensor cell line were developed as tools to perform this technique. This technique being simple and less expensive could significantly facilitate high throughput screening in calcium channel drug discovery.

Recombinant outer membrane protein 25c from Brucella abortus induces Th1 and Th2 mediated protection against Brucella abortus infection in mouse model.

Development of a safe and efficacious vaccine for brucellosis is a long standing challenge for scientists. Recognizing potential antigens towards developing vaccine candidate is crucial. Omp25c, a porin protein of Brucella, is a paralog of two previously identified promising vaccine candidates namely, Omp25 and Omp31, with notable sequence identity. Also, Omp25c is conserved in all major Brucella species. This highlights the possibility of employing this protein in multivalent subunit vaccine based approach of Brucella management. In this study, we were interested in examining the immunogenicity and protective efficacy of Omp25c against Brucella infections. Recombinant unlipidated form of this antigen (rOmp25c) produced, upon intraperitoneal immunization in BALB/c mice along with Freund's adjuvant, was confirmed to be highly immunogenic; leading to high IgG antibody titers during the study duration. The IgG2a/IgG2b ratio of anti-rOmp25c antibodies revealed elicitation of Th2 based humoral immunity. Lymphocyte proliferation study divulged induction of specific memory response and secretion of both Th1-type (IFN-γ, GM-CSF and TNF-α) and Th2-type cytokine (IL-5) from restimulated splenocytes of rOmp25c immunized mice. CD4 T-cell subpopulation was comparatively increased than total B cell subpopulation in case of immunized mice, indicating the induction of strong cell-mediated (Th1 biased) immunity than humoral (Th2) immunity. The collective Th1 plus Th2 immune response specific to rOmp25c could be the reason for protection against Brucella challenge observed in mice groups that was comparable with S19 vaccine strain. Thus, the study encourages rOmp25c as a potent candidate vaccine against brucellosis.

Evaluation of fimC and bdha based duplex PCR for specific identification and differentiation of Burkholderia pseudomallei from near-neighbor Burkholderia species.

Assays for the rapid detection and accurate differentiation of Burkholderia pseudomallei from near-neighbor species are urgently needed in melioidosis endemic regions due to the high associated mortality and biowarfare importance of the pathogen. PCR-based methods have revolutionized this field due to the accuracy, sensitivity, and specificity that are achievable in a rapid way. In this study, a compound molecular detection system, consisting of a duplex PCR assay, was developed for the specific identification of Burkholderia pseudomallei and differentiation from other Burkholderia species. For accurate identification of B. pseudomallei, we deciphered and adopted a novel gene termed putative fimbrial chaperone (fimC). d-beta hydroxybutyrate dehydrogenase (bdha), reported previously by our group for sequence-based differentiation of B. pseudomallei from other Burkholderia species, was employed as a genus-specific target. Enforcement of an internal amplification control in the PCR format ruled out possible false negative results in the assay. Thus, the developed PCR assay was highly specific (100%) in its detection features, and a clear detection sensitivity of 10 pg/µl for purified gDNA and 3 × 103 CFU/ml for B. pseudomallei spiked urine was recorded. Successful identification of B. pseudomallei from an experimental mouse model at both the genus and species level revealed the accurate diagnostic efficiency of the duplex PCR method.

Prevalence and Identification of Burkholderia pseudomallei and Near-Neighbor Species in the Malabar Coastal Region of India.

Accurate identification of pathogens with biowarfare importance requires detection tools that specifically differentiate them from near-neighbor species. Burkholderia pseudomallei, the causative agent of a fatal disease melioidosis, is one such biothreat agent whose differentiation from its near-neighbor species is always a challenge. This is because of its phenotypic similarity with other Burkholderia species which have a wide spread geographical distribution with shared environmental niches. Melioidosis is a major public health concern in endemic regions including Southeast Asia and northern Australia. In India, the disease is still considered to be emerging. Prevalence surveys of this saprophytic bacterium in environment are under-reported in the country. A major challenge in this case is the specific identification and differentiation of B. pseudomallei from the growing list of species of Burkholderia genus. The objectives of this study included examining the prevalence of B. pseudomallei and near-neighbor species in coastal region of South India and development of a novel detection tool for specific identification and differentiation of Burkholderia species. Briefly, we analyzed soil and water samples collected from Malabar coastal region of Kerala, South India for prevalence of B. pseudomallei. The presumptive Burkholderia isolates were identified using recA PCR assay. The recA PCR assay identified 22 of the total 40 presumptive isolates as Burkholderia strains (22.72% and 77.27% B. pseudomallei and non-pseudomallei Burkholderia respectively). In order to identify each isolate screened, we performed recA and 16S rDNA sequencing. This two genes sequencing revealed that the presumptive isolates included B. pseudomallei, non-pseudomallei Burkholderia as well as non-Burkholderia strains. Furthermore, a gene termed D-beta hydroxybutyrate dehydrogenase (bdha) was studied both in silico and in vitro for accurate detection of Burkholderia genus. The optimized bdha based PCR assay when evaluated on the Burkholderia isolates of this study, it was found to be highly specific (100%) in its detection feature and a clear detection sensitivity of 10 pg/µl of purified gDNA was recorded. Nucleotide sequence variations of bdha among interspecies, as per in silico analysis, ranged from 8 to 29% within the target stretch of 730 bp highlighting the potential utility of bdha sequencing method in specific detection of Burkholderia species. Further, sequencing of the 730 bp bdha PCR amplicon of each Burkholderia strain isolated could differentiate the species and the data was comparable with recA sequence data of the strains. All sequencing results obtained were submitted to NCBI database. Bayesian phylogenetic analysis of bdha in comparison with recA and 16S rDNA showed that the bdha gene provided comparable identification of Burkholderia species.

Differentiation of entC1 from entC2/entC3 with a single primer pair using simple and rapid SYBR Green-based RT-PCR melt curve analysis.

In spite of their involvement in foodborne illness, the epidemiological relevance of staphylococcal enterotoxin C (SEC) subtypes is poorly documented may be due to high sequence similarity. Among subtypes, SEC1, SEC2, and SEC3 exhibit more than 97 % homology because of which specific detection tools are seldom available to identify and differentiate them. In this study, a SYBR Green-based RT-PCR followed by melt curve analysis was developed for differentiation of entC1 from entC2/entC3 using a single primer pair. Nucleotide sequences of all three subtypes were analyzed using Clustal Omega program and the region with significant sequence variation/heterogeneity (where utmost SNPs were closely located and accessible for RT-PCR) was selected for amplification by designing a single primer pair that could amplify all three subtypes. In spite of same amplicon size, entC1 showed distinct melt peak at 76 °C. However, due to high similarity between entC2 and entC3, the developed format was deficient to discriminate between them and both showed melt peak at 82 °C. Reliability of developed RT-PCR was evaluated using various naturally contaminated samples and 91 food and clinical Staphylococcus aureus isolates where satisfactory results were obtained in comparison with commercial immunoassay kit and conventional PCRs using validated primers. To the best of our knowledge, this is the first method being reported to differentiate entC1 from entC2/entC3 using single primer pair which is unachievable by conventional PCR due to same amplicon size. As benefits, the method is sensitive, rapid, and inexpensive with no requirement of fluorescent probes, multiple primers, and post-PCR procedures. Thus, the assay might find its utility as a detection tool in epidemiological survey of foodborne outbreaks for simultaneous identification and differentiation of entC1 from entC2/entC3.

Evaluation of recombinant SEA-TSST fusion toxoid for protection against superantigen induced toxicity in mouse model.

Treatment of Staphylococcus aureus infections has become complicated owing to growing antibiotic resistance mechanisms and due to the multitude of virulence factors secreted by this organism. Failures with traditional monovalent vaccines or toxoids have brought a shift towards the use of multivalent formulas and neutralizing antibodies to combat and prevent range of staphylococcal infections. In this study, we evaluated the efficacy of a fusion protein (r-ET) comprising truncated regions of staphylococcal enterotoxin A (SEA) and toxic shock syndrome toxin (TSST-1) in generating neutralizing antibodies against superantigen induced toxicity in murine model. Serum antibodies showed specific reactivity to both SEA and TSST-1 native toxins. Hyperimmune serum from immunized animals protected cultured splenocytes from non-specific superantigen induced proliferation completely. Passive antibody administration prevented tissue damage from acute inflammation associated with superantigen challenge from S. aureus cell free culture supernatants. Approximately 80% and 50% of actively and passively immunized mice respectively were protected from lethal dose against S. aureus toxin challenge. This study revealed that r-ET protein is non-toxic and a strong immunogen which generated neutralizing antibodies and memory immune response against superantigen induced toxic effects in mice model.

Functional characterization and evaluation of in vitro protective efficacy of murine monoclonal antibodies BURK24 and BURK37 against Burkholderia pseudomallei.

Burkholderia pseudomallei, the causative agent of melioidosis has been recognized by CDC as a category B select agent. Although substantial efforts have been made for development of vaccine molecules against the pathogen, significant hurdles still remain. With no licensed vaccines available and high relapse rate of the disease, there is a pressing need for development of alternate protection strategies. Antibody-mediated passive protection is promising in this regard and our primary interest was to unravel this frontier of specific mAbs against Burkholderia pseudomallei infections, as functional characterization of antibodies is a pre-requisite to demonstrate them as protective molecules. To achieve this, we designed our study on in vitro-based approach and assessed two mAbs, namely BURK24 and BURK37, reactive with outer membrane proteins and lipopolysaccharide of the pathogen respectively, for their ability to manifest inhibitory effects on the pathogenesis mechanisms of B. pseudomallei including biofilm formation, invasion and induction of apoptosis. The experiments were performed using B. pseudomallei standard strain NCTC 10274 and a clinical isolate, B. pseudomallei 621 recovered from a septicemia patient with diabetic ailment. The growth kinetic studies of the pathogen in presence of various concentrations of each individual mAb revealed their anti-bacterial properties. Minimal inhibitory concentration and minimal bactericidal concentration of both the mAbs were determined by using standards of Clinical and Laboratory Standards Institute (CLSI) and experiments were performed using individual mAbs at their respective bacteriostatic concentration. As an outcome, both mAbs exhibited significant anti-Burkholderia pseudomallei properties. They limited the formation of biofilm by the bacterium and completely crippled its invasion into human alveolar adenocarcinoma epithelial cells. Also, the mAbs were appreciably successful in preventing the bacterium to induce apoptosis in A549 cells. The present study design revealed the protection attributes possessed by BURK24 and BURK37 that has to be further substantiated by additional in vivo studies.

Growth kinetics and immune response of chimeric foot-and-mouth disease virus serotype 'O' produced through replication competent mini genome of serotype Asia 1, 63/72, in BHK cell lines.

Regular vaccinations with potent vaccine, in endemic countries and vaccination to live in non-endemic countries are the methods available to control foot-and-mouth disease. Selection of candidate vaccine strain is not only cumbersome but the candidate should grow well for high potency vaccine preparation. Alternative strategy is to generate an infectious cDNA of a cell culture-adapted virus and use the replicon for development of tailor-made vaccines. We produced a chimeric 'O' virus in the backbone of Asia 1 and studied its characteristics. The chimeric virus showed high infectivity titre (>10(10)) in BHK 21 cell lines, revealed small plaque morphology and there was no cross reactivity with antiserum against Asia 1. The virus multiplies rapidly and reaches peak at 12h post infection. The vaccine prepared with this virus elicited high antibody titres.