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Abstract. In this paper, we define a one-vertex-extension tree for a distance-hereditary graph and show how to build it. We then give a unified approach to designing efficient dynamic programming algorithms for distance-hereditary graphs based upon the one-vertex-extension tree. We give linear time algorithms for the weighted vertex cover and weighted independent domination problems and give an O(n 2) time algorithm to compute a minimum fill-in and the treewidth for a distance-hereditary graph. IntroductionThe distance da (u, v) between two vertices u and v of a connected graph G is the minimum length of a u-v path in G. A graph is distance-hereditary if each pair of vertices are equidistant in every connected induced subgraph containing them. Distance-hereditary graphs form a subclass of perfect graphs [7,11,12] that are graphs G in which the maximum clique size equals the chromatic number for every induced subgraph of G. Properties and optimization problems in distance-hereditary graphs have been extensively studied during the past two decades [2,3, 4,7,8,9,11,12,17,19,20] which result in sequential algorithms that solve quite a few graph-theoretical problems on this special class of graphs. However, some problems remain unresolved on distance-hereditary graphs. For example, the independent domination problem and the vertex cover problem, the minimum fill-in and treewidth problem, and the edge domination problem etc. Most of the known polynomial time algorithms for distance-hereditary graphs utilize ad hoc techniques. In this paper, we first define a new data structure, called one-vertex-exlension tree, to represent a distance-hereditary graph in the form of a rooted tree. We give a new recursive definition for this class of graphs. We then show how to design dynamic programming algorithms based on this recursive definition. We present linear time algorithms to solve the weighted vertex cover problem and weighted independent domination problem and present a O(n 2) time algorithm to compute the minimum fill-in and treewidth for a distance-hereditary graph. The above three problems are NP-complete for general graphs [1,5,14], and the vertex cover problem is known to be MAX SNP-

BackgroundEnterovirus 71 (EV71) has caused several epidemics of hand, foot and mouth diseases (HFMD) in Asia and now is being recognized as an important neurotropic virus. Effective medications and prophylactic vaccine against EV71 infection are urgently needed. Based on the success of inactivated poliovirus vaccine, a prototype chemically inactivated EV71 vaccine candidate has been developed and currently in human phase 1 clinical trial.Principal FindingIn this report, we present the development of a serum-free cell-based EV71 vaccine. The optimization at each step of the manufacturing process was investigated, characterized and quantified. In the up-stream process development, different commercially available cell culture media either containing serum or serum-free was screened for cell growth and virus yield using the roller-bottle technology. VP-SFM serum-free medium was selected based on the Vero cell growth profile and EV71 virus production. After the up-stream processes (virus harvest, diafiltration and concentration), a combination of gel-filtration liquid chromatography and/or sucrose-gradient ultracentrifugation down-stream purification processes were investigated at a pilot scale of 40 liters each. Although the combination of chromatography and sucrose-gradient ultracentrifugation produced extremely pure EV71 infectious virus particles, the overall yield of vaccine was 7–10% as determined by a VP2-based quantitative ELISA. Using chromatography as the downstream purification, the virus yield was 30–43%. To retain the integrity of virus neutralization epitopes and the stability of the vaccine product, the best virus inactivation was found to be 0.025% formalin-treatment at 37°C for 3 to 6 days. Furthermore, the formalin-inactivated virion vaccine candidate was found to be stable for >18 months at 4°C and a microgram of viral proteins formulated with alum adjuvant could induce strong virus-neutralizing antibody responses in mice, rats, rabbits, and non-human primates.ConclusionThese results provide valuable information supporting the current cell-based serum-free EV71 vaccine candidate going into human Phase I clinical trials.

Enterovirus 71 (EV71) is now recognized as an emerging neurotropic virus in Asia and with Coxsackie virus (CV) it is the other major causative agent of hand-foot-mouth diseases (HFMD). Effective medications and/or prophylactic vaccines against HFMD are urgently needed. From a scientific (the feasibility of bioprocess, immunological responses and potency in animal challenge model) and business development (cost of goods) points of view, we in this review address and discuss the pros and cons of different EV71 vaccine candidates that have been produced and evaluated in animal models. Epitope-based synthetic peptide vaccine candidates containing residues 211–225 of VP1 formulated with Freund’s adjuvant (CFA/IFA) elicited low EV71 virus neutralizing antibody responses, but were protective in the suckling mouse challenge model. Among recombinant EV71 subunits (rVP1, rVP2 and rVP3) expressed in E. coli, purified and formulated with CFA/IFA, only VP1 elicited mouse antibody responses with measurable EV71-specific virus neutralization titers. Immunization of mice with either a DNA plasmid containing VP1 gene or VP1 expressed in Salmonella typhimurium also generated neutralizing antibody responses and protected animals against a live EV71 challenge. Recombinant EV71 virus-like particles (rVLP) produced from baculovirus formulated either with CFA/IFA or alum elicited good virus neutralization titers in both mice and non-human primates, and were found to be protective in the suckling mouse EV71 challenge model. Synthetic peptides or recombinant EV71 subunit vaccines (rVP1 and rVLP) formulated in alum were found to be poorly immunogenic in rabbits. Only formalin-inactivated (FI) EV71 virions formulated in alum elicited cross-neutralizing antibodies against different EV71 genotypes in mice, rabbits and non-human primates but induced weak neutralizing responses against CAV16. From a regulatory, economic and market acceptability standpoint, FI-EV71 virion vaccines are the most promising candidates and are currently being evaluated in human clinical trials. We further describe and analyze some new bioprocesses technologies that have great potential applications in EV71 vaccine development. This review also demonstrates the opportunities and challenges that the Asian vaccine industry faces today.

The immunity protein of colicin E7 (ImmE7) can bind specifically to the DNase-type colicin E7 and inhibit its bactericidal activity. Here we report the 1.8-A crystal structure of the ImmE7 protein. This is the first x-ray structure determined in the superfamily of colicin immunity proteins. The ImmE7 protein consists of four antiparallel ci-helices, folded in a topology similar to the architecture of a four-helix bundle structure. A region rich in acidic residues is identified. This negatively charged area has the greatest variability within the family of DNase-type immunity proteins; thus, it seems likely that this area is involved in specific binding to colicin. Based on structural, genetic, and kinetic data, we suggest that all the DNase-type immunity proteins, as well as colicins, share a "homologous-structural framework" and that specific interaction between a colicin and its cognate immunity protein relies upon how well these two proteins' charged residues match on the interaction surface, thus leading to specific immunity of the colicin.E-group colicins (from El to E9) are plasmid-borne antibioticlike bacteriocins that are active against sensitive Escherichia coli and closely related coliform bacteria (1). They bind to the vitamin B12 receptor, BtuB, and subsequently translocate across the outer and cytoplasmic membranes, inducing cell death (2). Three cytotoxic classes of E-group colicins have thus far been identified, including pore-forming colicins such as colicin El (3), RNase colicins such as colicins E3 (4) and E6 (5), and DNase colicins such as colicins E2 (6), E7 (7), E8 (8), and E9 (7). Colicin E7 (ColE7) is a nonspecific endonuclease that causes both single-and double-stranded breaks in the DNA of sensitive cells. Production of ColE7 is regulated by a "SOS" response operon that encodes ColE7, ImmE7 (immunity protein of colicin E7), and a lysis protein for transportation of the ColE7/ImmE7 complex. Immediately after production, colicin forms a complex with its coordinately produced ImmE and thus neutralizes its toxicity toward the host cell. The DNase-type colicins contain almost identical sequences in their translocation and receptor recognition domains, which are located in the N-terminal two-thirds of the sequence.C-terminal endonuclease domains (T2A domain) of DNasetype colicins are =80% identical, and sequences of their corresponding immunity proteins are 60-70% identical (7).However, despite the high sequence identities in either colicins or immunity proteins, an immunity protein can only completely protect a cell from the action of its own cognate colicin (9). Neither the mechanism for the specific protein-protein interaction between colicins and immunity proteins nor the inhibition of toxicity incurred after the formation of colicin/ ImmE complex has been explained yet. Thus the crystal structure of ImmE7 may provide invaluable information, expanding our limited knowledge of the specific interactions between proteins. Here we report the 1.8-A crystal structure of the ImmE7 protein...