Characterization and immunogenicity of a Shigella flexneri 2a O-antigen bioconjugate vaccine candidate.

Shigellosis remains a major cause of diarrheal disease in developing countries and causes substantial morbidity and mortality in children. Vaccination represents a promising preventive measure to fight the burden of the disease, but despite enormous efforts, an efficacious vaccine is not available to date. The use of an innovative biosynthetic Escherichia coli glycosylation system substantially simplifies the production of a multivalent conjugate vaccine to prevent shigellosis. This bioconjugation approach has been used to produce the Shigella dysenteriae type O1 conjugate that has been successfully tested in a phase I clinical study in humans. In this report, we describe a similar approach for the production of an additional serotype required for a broadly protective shigellosis vaccine candidate. The Shigella flexneri 2a O-polysaccharide is conjugated to introduced asparagine residues of the carrier protein exotoxin A (EPA) from Pseudomonas aeruginosa by co-expression with the PglB oligosaccharyltransferase. The bioconjugate was purified, characterized using physicochemical methods and subjected to preclinical evaluation in rats. The bioconjugate elicited functional antibodies as shown by a bactericidal assay for S. flexneri 2a. This study confirms the applicability of bioconjugation for the S. flexneri 2a O-antigen, which provides an intrinsic advantage over chemical conjugates due to the simplicity of a single production step and ease of characterization of the homogenous monomeric conjugate formed. In addition, it shows that bioconjugates are able to raise functional antibodies against the polysaccharide antigen.

Permian scorpions from the Petrified Forest of Chemnitz, Germany.

BACKGROUND: Paleozoic scorpions (Arachnida: Scorpiones) have been widely documented from the Carboniferous Period; which hosts a remarkable assemblage of more than sixty species including both putative stem- and crown-group fossils. By contrast the succeeding Permian Period is almost completely devoid of records, which are currently restricted to a trace fossil from the early Permian of New Mexico, USA and some limb fragments from the late Permian of the Vologda Region, Russia. RESULTS: ?Opsieobuthus tungeri sp. nov. from the Petrified Forest of Chemnitz, Germany represents the first complete body fossils of scorpions from the Permian. Explosive volcanism preserved these remarkable specimens in situ as part of the palaeosol horizon and bedrock of the Petrified Forest, immediately beneath the Zeisigwald tuff horizon. This dates to the early Permian (Sakmarian) or ca. 291 Ma. Intriguingly, the specimens were obtained from a palaeosol horizon with a compacted network of different-sized woody roots and thus have been preserved in situ in their likely life position, even within their original burrows. Differences in the structure of the comb-like pectines in the two fossils offer evidence for sexual dimorphism, and permit further inferences about the ecology and perhaps even the reproductive biology of these animals. CONCLUSIONS: As putative members of a Coal Measures genus, these fossils suggest that at least some Carboniferous scorpion lineages extended their range further into the Permian. This contributes towards a picture of scorpion evolution in which both basal and derived (orthostern) forms coexisted for quite some time; probably from the end of the Carboniferous through to at least the mid Triassic.

Arthroscopic Repair of Massive Cuff Tears With Large Subscapularis Tendon Ruptures (Lafosse III/IV): A Prospective Magnetic Resonance Imaging-Controlled Case Series of 26 Cases With a Minimum Follow-up of 1 Year.

PURPOSE: To prospectively assess arthroscopic repair of massive cuff tears (MCT) in a highly selective patient group with large subscapularis (SSC) tendon tears by means of clinical results and magnetic resonance imaging (MRI) studies. METHODS: Between April 2009 and December 2010, 26 patients with MCT were treated with arthroscopic rotator cuff repair. Only lesions involving a large tear of the SSC tendon (Lafosse III or IV) in combination with a complete tear of the supraspinatus (SSP) tendon and a tear of at least the anterior third of the infraspinatus (ISP) tendon were included. Minimum follow-up was 1 year. Pre- and postoperative assessment included a standardized clinical examination, subjective patient outcome, and MRI (structural integrity, fatty muscle infiltration, and muscular mass). RESULTS: Mean follow-up was 17 months (range, 12 to 34 months). MRI was performed in 25 patients. In 21 (84%) the cuff repair was intact. A partial retear of the SSC was found in 2 patients (8%). In 2 patients (8%) a full-thickness retear of the posterosuperior cuff was observed (1 SSP, 1 SSP/ISP). A significant increase of the muscle mass and decrease of fatty infiltration was observed for the SSC and SSP but not for the ISP. The mean Constant-Murley score improved from 36 to 86 points (P < .001) with all its subscores as well (P < .001). Muscular strength improved for the SSC (4.9 v 3.0, P < .001), SSP (4.6 v 2.9, P < .001), and ISP (4.8 v 3.4, P < .001). Overall patient satisfaction was high (3.6 ± 0.8). CONCLUSIONS: Arthroscopic repair of MCT involving the ISP, SSP, and large tears of the SSC provides a reliable tendon healing, in particular for the SSC tendon, combined with good functional results. LEVEL OF EVIDENCE: Level IV, therapeutic case series.

Arthroscopic repair of traumatic isolated subscapularis tendon lesions (Lafosse Type III or IV): a prospective magnetic resonance imaging-controlled case series with 1 year of follow-up.

PURPOSE: The purpose of this study was to prospectively assess the efficacy of arthroscopic repair of isolated high-grade subscapularis (SSC) tendon lesions by means of clinical follow-up combined with magnetic resonance imaging investigations. METHODS: Between January 2008 and September 2010, 11 patients (9 men and 2 women; mean age, 45 ± 10 years) with Lafosse type III or IV traumatic isolated SSC tendon lesions underwent arthroscopic repair including tenodesis of the long head of the biceps tendon. All patients were preoperatively assessed by clinical examination (Constant-Murley score [CMS]) and contrast-enhanced magnetic resonance arthrography. At 1 year of follow-up, specific clinical SSC tests, the CMS, and the loss of external rotation were evaluated. A native magnetic resonance investigation was performed to assess the structural integrity of the repair. The SSC muscle was compared with its preoperative condition regarding fatty infiltration and size (cross-sectional area). Patient satisfaction was graded from 1 (poor) to 4 (excellent). RESULTS: The mean time interval from trauma to surgery was 3.7 months. A concomitant lesion of the biceps tendon was observed in 10 patients (91%). The mean CMS improved from 44 to 89 points (P < .001). The functional tests showed a significant increase in strength (P < .05) (belly-press test, 4.8 v 2.9; lift-off test, 4.8 v 2.9). The mean loss of external rotation at 0° of abduction was 10° compared with the contralateral side (P < .05). Patient satisfaction was high. Magnetic resonance imaging evaluation showed complete structural integrity of the tendon repair in all studies. The SSC showed a significant decrease in fatty infiltration and increase in the cross-sectional area. CONCLUSIONS: Arthroscopic repair of higher-grade isolated SSC lesions provides reliable tendon healing accompanied by excellent functional results 1 year after surgery. LEVEL OF EVIDENCE: Level IV, prospective therapeutic case series.

Immunization of Rabbits with a Quadrivalent Shigella Bioconjugate Vaccine Induces Functional Antibodies Reactive with Shigella Isolates from Kenya.

Diarrheal diseases are a leading cause of global morbidity and mortality, disproportionately affecting children in resource-limited settings. Although improvements in hygiene and access to clean water are helpful, vaccines are considered essential due to the low infectious dose of Shigella species and increasing antibiotic resistance. Building on achievements with conjugate vaccines, a safe and immunogenic novel bioconjugate vaccine linking Shigella O-antigen to Pseudomonas aeruginosa exoprotein A has been developed to induce immunity against Shigella flexneri 2a, 3a, and 6 and S. sonnei. This study evaluated the breadth of reactivity and functionality of pooled serum from rabbits immunized with monovalent and quadrivalent Shigella bioconjugates formulated with or without an adjuvant against Shigella serotypes isolated in Kenya. Rabbit sera were assayed by colony blot for reactivity with 67 isolates of Shigella serotypes targeted by the vaccine, S. flexneri (2a, 3a, and 6) and S. sonnei, and 42 isolates of Shigella serotypes not targeted by the vaccine, S. flexneri (1b, 2b, 4a, and 4b), S. boydii, and S. dysenteriae. Shigella isolates testing positive in the colony blot assay were then used to assess functional activity using a bactericidal assay. Of the 41 Shigella isolates targeted by the vaccine, 22 were reactive with the adjuvanted quadrivalent and the respective monovalent rabbit sera. The S. flexneri 2a and 3a monovalent rabbit serum cross-reacted with S. flexneri 3a, 2b, and 2a, respectively. Immunization with the adjuvanted quadrivalent vaccine also induced cross-reactivity with isolates of S. flexneri 2b, 4a, and 4b. Collectively, these results suggest that the Shigella quadrivalent vaccine may be more broadly protective than designed, offering a promising solution to Shigella infections. IMPORTANCE Diarrheal diseases are the third leading cause of death globally, disproportionally affecting low- to middle-income countries like Kenya, with Shigella species being the leading cause of bacterial diarrhea, especially in children. The low infectious dose and high antibiotic resistance levels complicate treatment, leading to long-term sequelae that necessitate control measures such as vaccines to reduce morbidity and mortality rates, especially among children under 5 years of age. A quadrivalent bioconjugate Shigella vaccine was recently developed to safely and effectively induce immunity against four important Shigella spp. This study demonstrates the breadth of reactivity and functionality of the parenterally administered bioconjugate vaccine by evaluating the ability of rabbit sera to bind and kill Shigella isolates recently collected in Kenya. These results suggest that the Shigella quadrivalent vaccine may be more broadly protective than designed and may offer a promising solution to the morbidity and mortality associated with Shigella infections.

A study on structural, optical, and electrical characteristics of perovskite CsPbBr3 QD/2D-TiSe2 nanosheet based nanocomposites for optoelectronic applications.

Lead halide perovskite (CsPbBr3) quantum dots (QDs) and two-dimensional (2D) layered transition metal dichalcogenides have a significant application in solution-processed optoelectronic devices. Here, we report the oleylamine-assisted exfoliation of TiSe2 nanosheets (NSs) in dichlorobenzene with high concentration and stable dispersion. The functionalized TiSe2 NSs were used to synthesize the solution-processed perovskite CsPbBr3 QD/TiSe2 NS-based nanocomposite. The perovskite QDs and TiSe2 NSs were characterized by different techniques. The strong photoluminescence (PL) quenching and decreased lifetime decay of the nanocomposite indicates efficient charge transfer from photo-excited CsPbBr3 to TiSe2 NSs. The calculated charge-transfer rate constant (KET) from photo-excited CsPbBr3 to TiSe2 NSs increased from 1.50 × 108 to 2.79 × 108 s-1 in different concentrations of TiSe2 NSs (5 to 20 µg mL-1), respectively. Furthermore, the photo-currents of CsPbBr3 QD/TiSe2 NS nanocomposite devices were dramatically enhanced ∼2 times compared to pristine CsPbBr3 QD based devices, which supports the charge transfer and charge separation in nanocomposite devices.

Enhanced T-cell immunogenicity of plasmid DNA vaccines boosted by recombinant modified vaccinia virus Ankara in humans.

In animals, effective immune responses against malignancies and against several infectious pathogens, including malaria, are mediated by T cells. Here we show that a heterologous prime-boost vaccination regime of DNA either intramuscularly or epidermally, followed by intradermal recombinant modified vaccinia virus Ankara (MVA), induces high frequencies of interferon (IFN)-gamma-secreting, antigen-specific T-cell responses in humans to a pre-erythrocytic malaria antigen, thrombospondin-related adhesion protein (TRAP). These responses are five- to tenfold higher than the T-cell responses induced by the DNA vaccine or recombinant MVA vaccine alone, and produce partial protection manifest as delayed parasitemia after sporozoite challenge with a different strain of Plasmodium falciparum. Such heterologous prime-boost immunization approaches may provide a basis for preventative and therapeutic vaccination in humans.

A study on chemical exfoliation and structural and optical properties of two-dimensional layered titanium diselenide.

Titanium diselenide (TiSe2) is the least studied member of the transition metal dichalcogenide family due to a lack of available synthesis methodology, controlled bandgap engineering, and rapid characterization of layers. In this paper, we report the chemical exfoliation of TiSe2 platelets synthesized by the chemical vapor transport route in ortho-dichlorobenzene (o-DCB) functionalized with oleylamine (OLA), for the first time to the best of our knowledge. It is found that the addition of OLA supports the formation of a stable dispersion of a large area of the TiSe2 sheets due to surface capping with the OLA molecules indicating the importance of the ligand in dispersion behavior. The X-ray diffraction pattern confirms the hexagonal structure of the TiSe2 platelets with the space group P3[combining macron]m1 while Raman spectroscopy reveals that two modes of vibration i.e. A1g and E2g exist with layered structures having dimensions in micrometers as confirmed by scanning electron microscopy. Fourier transform infrared spectroscopy confirms the successful functionalization of chemically exfoliated TiSe2 nanosheets. Field-emission scanning electron microscopy reveals that exfoliated TiSe2 has a thickness of 15-55 nm whereas high-resolution transmission electron microscopy indicates thicker sheets for ligand-free exfoliated TiSe2 which are crystalline. Atomic force microscopy confirms the formation of nanosheets. UV-Visible, photoluminescence, and time-resolved PL spectroscopy showed an enhanced effect and better average lifetime of excitation for the exfoliated sheets with OLA than those without OLA. The C-V studies reveal that with increasing scan rate, the corresponding current also increases. The present study offers the possibility of their utilization in optoelectronics, advanced low-power electronics, voltage-controlled oscillators, ultra-fast electronics, and electrochemical devices.

Clinical and immunological responses in metastatic melanoma patients vaccinated with a high-dose poly-epitope vaccine.

BACKGROUND: Safety and cellular immunogenicity of rising doses and varying regimens of a poly-epitope vaccine were evaluated in advanced metastatic melanoma. The vaccine comprised plasmid DNA and recombinant modified vaccinia virus Ankara (MVA) both expressing a string (Mel3) of seven HLA.A2/A1 epitopes from five melanoma antigens. METHODS: Forty-one HLA-A2 positive patients with stage III/IV melanoma were enrolled. Patient groups received one or two doses of DNA.Mel3 followed by escalating doses of MVA.Mel3. Immunisations then continued eight weekly in the absence of disease progression. Epitope-specific CD8+ T cell responses were evaluated using ex-vivo tetramer and IFN-gamma ELISPOT assays. Safety and clinical responses were monitored. RESULTS: Prime-boost DNA/MVA induced Melan-A-specific CD8+ T cell responses in 22/31 (71%) patients detected by tetramer assay. ELISPOT detected a response to at least one epitope in 10/31 (32%) patients. T cell responder rates were <50% with low-dose DNA/MVA, or MVA alone, rising to 91% with high-dose DNA/MVA. Among eight patients showing evidence of clinical benefit-one PR (24 months+), five SD (5 months+) and two mixed responses-seven had associated immune responses. Melan-A-tetramer+ immunity was associated with a median 8-week increase in time-to-progression (P = 0.037) and 71 week increase in survival (P = 0.0002) compared to non-immunity. High-dose vaccine was well tolerated. The only significant toxicities were flu-like symptoms and injection-site reactions. CONCLUSIONS: DNA.Mel3 and MVA.Mel3 in a prime-boost protocol generated high rates of immune response to melanoma antigen epitopes. The treatment was well tolerated and the correlation of immune responses with patient outcomes encourages further investigation.

Tuning the performance of vanadium redox flow batteries by modifying the structural defects of the carbon felt electrode.

Polyacrylonitrile (PAN)-based carbon felt was subjected to N2-plasma treatment to increase the heteroatom defects and reactive edge sites as a method to increase the performance in vanadium redox flow batteries (VRFBs). N-doping in the felt was mainly in the form of pyrrolic and pyridinic nitrogen. Even though the amount of oxygen functional groups on the N2-plasma-treated sample was very low, the felt showed enhanced electrochemical performance for both V3+/V2+ as well as V5+/V4+ redox reactions. The result is highly significant as the pristine electrode with the same amount of oxygen functional groups showed significantly less activity for the V3+/V2+ redox reaction. Overall, the single-flow cell experiments with N2-plasma-treated felt showed superior performance compared to the pristine sample. Therefore, the enhanced performance observed for the N2-plasma-treated sample should be attributed to the increase in defects and edge sites. Thus, from the present study, it can be concluded that an alternate way to increase the performance of the VRFBs is to introduce specific defects such as N-doping/substitution or to increase the edge sites. In other words, defects induced in the carbon felt such as heteroatom doping are as beneficial as the presence of oxygen functional groups for the improved performance of VRFBs. Therefore, for an optimum performance of VRFBs, defects such as N-substitution as well as oxygen functionality should be tuned.

Polysaccharide conjugate vaccine protein carriers as a "neglected valency" - Potential and limitations.

The development of vaccines against polysaccharide-encapsulated pathogens (e.g. Haemophilus influenzae type b, pneumococci, meningococci) is challenging because polysaccharides do not elicit a strong and long-lasting immune response (i.e. T-cell independent). This can be overcome by conjugating the polysaccharide to a protein carrier (e.g. tetanus toxoid, cross-reacting material 197 [CRM]), which vastly improves the immune response and induces memory to the polysaccharide (T-cell dependent). Although it is well documented that protein carriers additionally induce an immune response against themselves, this potential "additional valency" has so far not been recognized. The only exception is for the protein D carrier (derived from non-typeable Haemophilus influenzae [NTHi]) used in a pneumococcal conjugate vaccine, which may have a beneficial impact on NTHi acute otitis media. In this review, we describe the immunogenicity of various protein carriers and discuss their potential dual function: as providers of T-cell helper epitopes and as protective antigens. If this "additional valency" could be proven to be protective, it may be possible to consider its potential effect on the number of required immunizations. We also describe the potential for positive or negative interference between conjugate vaccines using the same protein carriers, the resulting desire for novel carriers, and information on potential new carriers. The range of conjugate vaccines is ever expanding, with different carriers and methods of conjugation. We propose that new conjugate vaccine trials should assess immunogenicity to both the polysaccharide and carrier. Ultimately, this so-far "neglected valency" could be an exploitable characteristic of polysaccharide conjugate vaccines.

In Vivo Testing of Extracorporeal Membrane Ventilators: iLA-Activve Versus Prototype I-Lung.

A side-by-side comparison of the decarboxylation efficacy of two pump-driven venovenous extracorporeal lung assist devices, i.e., a first prototype of the new miniaturized ambulatory extracorporeal membrane ventilator, I-lung versus the commercial system iLA-activve for more than a period of 72 hours in a large animal model. Fifteen German Landrace pigs were anesthetized and underwent mechanical hypoventilation to induce severe hypercapnia. Decarboxylation was accomplished by either the I-lung or the iLA-activve via a double lumen catheter in the jugular vein. Sham-operated pigs were not connected to extracorporeal devices. Cardiovascular, respiratory, and metabolic parameters were continuously monitored, combined with periodic arterial blood sampling for subsequent clinical blood diagnostics, such as gas exchange, hemolysis, coagulation parameters, and cytokine profiles. At the termination of the studies, lung tissue was harvested and examined histologically for pulmonary morphology and leukocyte tissue infiltration. Both extracorporeal devices showed high and comparable efficacy with respect to carbon dioxide elimination for more than 72 hours and were not associated with either bleeding events or clotting disorders. Pigs of both groups showed cardiovascular and hemodynamic stability without marked differences to sham-operated animals. Groups also did not differ in terms of inflammatory and metabolic parameters. We established a preclinical in vivo porcine model for comparative long-term testing of I-lung and iLA-activve. The I-lung prototype proved to be safe and feasible, providing adequate decarboxylation without any adverse events. Once translated into the clinical treatment, the new miniaturized and transportable I-lung device might represent a promising tool for treating awake and mobilized patients with decompensated pulmonary disorders.

Immunogenicity and safety of a tetravalent E. coli O-antigen bioconjugate vaccine in animal models.

BACKGROUND: Extra-intestinal pathogenic Escherichia coli (ExPEC) are major human pathogens; however, no protective vaccine is currently available. We assessed in animal models the immunogenicity and safety of a 4-valent E. coli conjugate vaccine (ExPEC-4V, serotypes O1, O2, O6 and O25 conjugated to Exotoxin A from Pseudomonas aeruginosa (EPA)) produced using a novel in vivo bioconjugation method. METHODS: Three doses of ExPEC-4V (with or without aluminum hydroxide) were administered to rabbits (2µg or 20µg per O-antigen, subcutaneously), mice (0.2µg or 2µg per O-antigen, subcutaneously) and rats (0.4µg or 4µg per O-antigen, intramuscularly). Antibody persistence and boostability were evaluated in rats using O6-EPA monovalent conjugate (0.4µg O-antigen/dose, intramuscularly). Toxicity was assessed in rats (16µg total polysaccharide, intramuscularly). Serum IgG and IgM antibodies were measured by ELISA. RESULTS: Robust antigen-specific IgG responses were observed in all animal models, with increased responses in rabbits when administered with adjuvant. O antigen-specific antibody responses persisted up to 168days post-priming. Booster immunization induced a rapid recall response. Toxicity of ExPEC-4V when administered to rats was considered to be at the no observed adverse effect level. CONCLUSIONS: ExPEC-4V conjugate vaccine showed good immunogenicity and tolerability in animal models supporting progression to clinical evaluation.

Progress toward a universal H5N1 vaccine: a recombinant modified vaccinia virus Ankara-expressing trivalent hemagglutinin vaccine.

BACKGROUND: The rapid evolution of new sublineages of H5N1 influenza poses the greatest challenge in control of H5N1 infection by currently existing vaccines. To overcome this, an MVAtor vector expressing three H5HA antigens A/Vietnam/1203/04, A/Indonesia/669/06 and A/Anhui/01/05 (MVAtor-tri-HA vector) was developed to elicit broad cross-protection against diverse clades by covering amino acid variations in the major neutralizing epitopes of HA among H5N1 subtypes. METHODS: BALB/c mice and guinea pigs were immunized i.m. with 8×107 TCID50/animal of MVAtor-tri-HA vector. The immunogenicity and cross-protective immunity of the MVAtor-tri-HA vector was evaluated against diverse clades of H5N1 strains. RESULTS: The results showed that mice immunized with MVAtor-tri-HA vector induced robust cross-neutralizing immunity to diverse H5N1 clades. In addition, the MVAtor-tri-HA vector completely protected against 10 MLD50 of a divergent clade of H5N1 infection (clade 7). Importantly, the serological surveillance of post-vaccinated guinea pig sera demonstrated that MVAtor-tri-HA vector was able to elicit strong cross-clade neutralizing immunity against twenty different H5N1 strains from six clades that emerged between 1997 and 2012. CONCLUSIONS: The present findings revealed that incorporation of carefully selected HA genes from divergent H5N1 strains within a single vector could be an effective approach in developing a vaccine with broad coverage to prevent infection during a pandemic situation.

Matriderm versus Integra: a comparative experimental study.

AIM: To compare engraftment rates and vascularisation in a rat model using either Integra Artificial Skin or Matriderm. METHODS: Matriderm and the dermal part of Integra were compared in a two-step procedure including matrix implantation and subsequent epidermal grafting. Neonatal rat epidermis was used as coverage to test for rapid and complete take. RESULTS: Efficiency and quality of vascularisation expressed by take rate of epidermis, and thickness of resulting neodermis, were identical for both matrices. CONCLUSION: This first comparison of Matriderm with Integra in a rat model revealed no major differences in engraftment rates or vascularisation.

Plasmid DNA and viral vector-based vaccines for the treatment of cancer.

Plasmid DNA and viral vector-based cancer vaccines have many inherent features that make them promising cancer vaccine candidates. This review focuses on the use of plasmid DNA and viral vector vaccines to deliver tumour-specific antigens to induce a tumour-specific immune response. Examples of different antigen delivery systems that have been tested in recent clinical trials are summarised and advantages and disadvantages of a number of delivery systems and approaches are discussed. Finally, an outlook on how plasmid DNA and viral vectors might be developed further as cancer vaccines is provided.

Treating Displaced Distal Forearm Fractures in Children.

PURPOSE: Distal forearm fractures are among the most common fractures in children. In the past few years the option of percutaneous pinning has gained more attention in the treatment of unstable fractures. However, it remains unclear in which cases a fracture or its reduction should be considered unstable. STUDY DESIGN: In order to evaluate which type of fractures profit most from additional pinning after closed reduction, we performed a retrospective analysis of 225 consecutive cases using the recently published AO pediatric classification of long bone fractures. RESULTS: After closed reduction, position in the cast was lost in 23% of the cases. The proportion of unstable reductions was much higher in completely displaced fractures. The amount of dislocation was more important than the type of fracture according to the AO classification proposal. CONCLUSIONS: Fully displaced fractures should always be reduced in a setting with pins immediately available. If anatomical reduction cannot be achieved, pinning is advocated. The AO proposal for pediatric long bone fracture classification could be a useful tool to render the diverse studies more comparable. However, the important feature of complete versus subtotal displacement is lacking.

A clinical trial of prime-boost immunisation with the candidate malaria vaccines RTS,S/AS02A and MVA-CS.

Heterologous prime-boost immunisation with RTS,S/AS02A and the poxvirus MVA-CS was evaluated in 18 healthy malaria-naïve subjects in Oxford. Both priming with RTS,S and boosting MVA-CS, and the reverse, were found to be safe and well tolerated. T cell responses as measured by IFN-gamma ex vivo ELISPOT were induced, but the responses were low to moderate in both groups, with heterologous boosting yielding only small increments in T cell immunogenicity and no increased antibody response. Protection against 3D7 Plasmodium falciparum sporozoite challenge 4 weeks after the final vaccination was equal for both regimens at 33% (95% C.I. 4.3-77.7%), with one subject remaining fully protected on rechallenge at 5 months.

Recombinant modified vaccinia Ankara primes functionally activated CTL specific for a melanoma tumor antigen epitope in melanoma patients with a high risk of disease recurrence.

Recombinant plasmid DNA and attenuated poxviruses are under development as cancer and infectious disease vaccines. We present the results of a phase I clinical trial of recombinant plasmid DNA and modified vaccinia Ankara (MVA), both encoding 7 melanoma tumor antigen cytotoxic T lymphocyte (CTL) epitopes. HLA-A*0201-positive patients with surgically treated melanoma received either a "prime-boost" DNA/MVA or a homologous MVA-only regimen. Ex vivo tetramer analysis, performed at multiple time points, provided detailed kinetics of vaccine-driven CTL responses specific for the high-affinity melan-A(26-35) analogue epitope. Melan-A26-35-specific CTL were generated in 2/6 patients who received DNA/MVA (detectable only after the first MVA injection) and 4/7 patients who received MVA only. Ex vivo ELISPOT analysis and in vitro proliferation assays confirmed the effector function of these CTL. Responses were seen in smallpox-vaccinated as well as vaccinia-naive patients, as defined by anti-vaccinia antibody responses demonstrated by ELISA assay. The observations that 1) CTL responses were generated to only 1 of the recombinant epitopes and 2) that the magnitude of these responses (0.029-0.19% CD8(+) T cells) was below the levels usually seen in acute viral infections suggest that to ensure high numbers of CTL specific for multiple recombinant epitopes, a deeper understanding of the interplay between CTL responses specific for the viral vector and recombinant epitopes is required.

Neutron diffraction study of [H(4)Co(4)(C(5)Me(4)Et)(4)], a tetrahedral metal cluster complex with four face-bridging hydride ligands.

A single-crystal neutron diffraction analysis of the cluster complex [H(4)Co(4)(C(5)Me(4)Et)(4)] was carried out on the new quasi-Laue diffractometer VIVALDI at the Institut Laue-Langevin. The structure consists of four face-bridging hydrides attached to a tetrahedral cobalt metal core. Average distances and angles in the core of the molecule are as follows: Co-Co = 2.571(8), Co-C = 2.158(6), Co-H = 1.749(7), H.H = 2.366(9) A; Co-H-Co = 94.6(3), H-Co-H = 85.1(3) degrees. The hydride ligands are located off the Co-Co-Co planes by an average distance of 0.923(8) A. It is suggested that the dimensions of the HCo(3) fragments found in this molecule provide reasonable estimates for analogous distances and angles associated with chemisorbed H atoms situated on the 3-fold hollows of a cobalt surface. Crystallographic details: space group P2(1)/a (monoclinic); a = 21.979(2), b = 10.924(1), c = 34.406(2) A; beta = 90.81(1) degrees; Z = 8. Final agreement factor: R(F) = 0.099 for 3779 reflections [I > 2sigma(I)] collected at 20 K.