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Traditional blood sampling by venipuncture is cumbersome and relatively expensive. Dried blood spot (DBS) sampling is desirable because of its ease of sample collection, transportation and storage. It has been used in clinical diagnosis but not been thoroughly studied for the potential use to assess the immune status of individuals following natural infection or preventive vaccination. The goal of this study was to compare DBS to traditional blood samplings in detection of antibodies in individuals vaccinated against measles, hepatitis A, tetanus, influenza and varicella zoster. Enzyme linked immunosorbent assay (ELISA) was used to test DBS eluates and serum samples for antibodies against measles, varicella, tetanus and hepatitis A. Sensitivities, specificities, and correlation coefficients were evaluated to compare optical density (OD) values of paired serum and DBS samples. The long-term stability of DBS samples at different temperatures was assessed using simulated immune measles blood. DBS OD was highly correlated with serum OD for antibodies to measles (r = 0.93), varicella (r = 0.82), and tetanus (r = 0.91). Sensitivities of DBS OD ranged from 86-99% and specificities ranged from 96-100% using cut-offs established by each assay. By contrast, the hepatitis A data showed a low sensitivity (31%) and weak correlation (r = 0.14) between DBS and serum samples. Antibody titers in serum samples for anti-influenza A (H1N1 and H3N1) failed to correlate in DBS eluates in HAI and MN assays. DBS samples were stable for 4 weeks when stored at room temperature and for 6 months at 4°C. DBS sampling was sensitive, specific, and highly correlated with traditional venipuncture sampling in detection of antibodies against measles, tetanus and varicella zoster, but not hepatitis A and influenza. Thus, the success of using DBS sampling to assess the antibody levels in immunized individuals may be dependent on the pathogens and the development of the assay used., Competing Interests: The authors have declared that no competing interests exist.

We conducted an open label, dose escalation Phase 1 clinical trial of a tetravalent dengue DNA vaccine (TVDV) formulated in Vaxfectin ® to assess safety and immunogenicity. A total of 40 dengue- and flavivirus-naive volunteers received either low-dose (1 mg) TVDV alone ( N = 10, group 1), low-dose TVDV (1 mg) formulated in Vaxfectin ( N = 10, group 2), or high-dose TVDV (2 mg, group 3) formulated in Vaxfectin ® ( N = 20). Subjects were immunized intramuscularly with three doses on a 0-, 30-, 90-day schedule and monitored. Blood samples were obtained after each immunization and various time points thereafter to assess anti-dengue antibody and interferon gamma (IFNγ) T-cell immune responses. The most common adverse events (AEs) across all groups included mild to moderate pain and tenderness at the injection site, which typically resolved within 7 days. Common solicited signs and symptoms included fatigue (42.5%), headache (45%), and myalgias (47.5%). There were no serious AEs related to the vaccine or study procedures. No anti-dengue antibody responses were detected in group 1 subjects who received all three immunizations. There were minimal enzyme-linked immunosorbent assay and neutralizing antibody responses among groups 2 and 3 subjects who completed the immunization schedule. By contrast, IFNγ T-cell responses, regardless of serotype specificity, occurred in 70%, 50%, and 79% of subjects in groups 1, 2, and 3, respectively. The largest IFNγ T-cell responses were among group 3 subjects. We conclude that TVDV was safe and well-tolerated and elicited predominately anti-dengue T-cell IFNγ responses in a dose-related fashion.

We describe the results from a human clinical trial of a dengue virus serotype-1, purified-inactivated vaccine (DENV-1 PIV) adjuvanted with aluminum hydroxide. This first-in-man, Phase 1, open-label clinical trial consisted of two groups of flavivirus-naïve healthy adult volunteers that received two intramuscular vaccine doses of either 2.5 μg or 5 μg of DENV-1 PIV administered on days 0 and 28. Following vaccination, both vaccine doses exhibited an acceptable safety profile with minimal injection site and systemic reactions. By study day 42, 2 weeks following the second vaccine dose, all volunteers in both vaccine groups developed serum-neutralizing antibodies against DENV-1. Additional testing using an enzyme-linked immunosorbent assay demonstrated induction of a humoral immune response following both vaccine doses. The DENV-1 PIV was safe and immunogenic in a small number of volunteers supporting development and further testing of a tetravalent DENV PIV formulation., (© The American Society of Tropical Medicine and Hygiene.)

Influenza-specific hemaggluitination inhibition (HAI) antibody titer, an indicator of immunity to influenza, is often used to measure exposure to influenza in surveillance and immunogenicity studies. Traditionally, serum has been the specimen of choice for HAI assays, but a desire to reduce the amount of blood collected during studies and the availability of plasma in archived sample collections warrant the evaluation of plasma for HAI titer. Therefore, the relationship between serum and plasma HAI titer values is of great interest. Here, we compare HAI titers determined on temporally matched serum and plasma (citrated and heparinized) using influenza A and B viruses. Bland-Altman plots, McNemar's test, and geometric coefficient of variation were used respectively for evaluating agreement, correlation and variability in the serum-plasma titer results. We observed a high degree of agreement (80.5%-98.8%) and correlation (r = 0.796-0.964) in the serum and matched plasma titer values although plasma titers were generally lower than corresponding serum titers. Calculated seropositive (HAI ≥40) rates were higher using serum titers than with plasma titers, but seroconversion rates were unaffected by sample type. Stronger agreement and decreased variability in titers were seen between serum and citrated plasma than between serum and heparinized plasma. Overall, these data suggest that serum or plasma can be used in serodiagnostic HAI assays, but seropositive rates may be underestimated using plasma HAI titers. The type of anticoagulant present in plasma may affect HAI titer values and warrants further investigation.

Vaccination with plasmid DNA against infectious pathogens including dengue is an active area of investigation. By design, DNA vaccines are able to elicit both antibody responses and cellular immune responses capable of mediating long-term protection. Great technical improvements have been made in dengue DNA vaccine constructs and trials are underway to study these in the clinic. The scope of this review is to highlight the rich history of this vaccine platform and the work in dengue DNA vaccines accomplished by scientists at the Naval Medical Research Center. This work resulted in the only dengue DNA vaccine tested in a clinical trial to date. Additional advancements paving the road ahead in dengue DNA vaccine development are also discussed., (Published by Elsevier Ltd.)

Candidate dengue DNA vaccine constructs for each dengue serotype were developed by incorporating pre-membrane and envelope genes into a plasmid vector. A Phase 1 clinical trial was performed using the dengue virus serotype-1 (DENV-1) vaccine construct (D1ME(100)). The study was an open-label, dose-escalation, safety and immunogenicity trial involving 22 healthy flavivirus-naïve adults assigned to one of two groups. Each group received three intramuscular injections (0, 1, and 5 months) of either a high dose (5.0mg, n=12) or a low dose (1.0mg, n=10) DNA vaccine using the needle-free Biojector(®) 2000. The most commonly reported solicited signs and symptoms were local mild pain or tenderness (10/22, 45%), local mild swelling (6/22, 27%), muscle pain (6/22, 27%) and fatigue (6/22, 27%). Five subjects (41.6%) in the high dose group and none in the low dose group developed detectable anti-dengue neutralizing antibodies. T-cell IFN gamma responses were detected in 50% (4/8) and 83.3% (10/12) of subjects in the low and high dose groups, respectively. The safety profile of the DENV-1 DNA vaccine is acceptable at both doses administered in the study. These results demonstrate a favorable reactogenicity and safety profile of the first in human evaluation of a DENV-1 DNA vaccine., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Tumor necrosis factor-alpha inhibitors are important adjunctive therapies for rheumatologic diseases. These agents increase the risk for granulomatous disease. We present a case of a woman with severe rheumatoid arthritis on infliximab who developed multiple nodular skin lesions. Biopsies grew Mycobacterium marinum. New lesions developed through therapy, necessitating surgical debulking.

Objective: To describe risks for, and microbiology and antimicrobial resistance patterns of, war trauma associated infections from Operation Iraqi Freedom., Background: : The invasion of Iraq resulted in casualties from high-velocity gunshot, shrapnel, and blunt trauma injuries as well as burns. Infectious complications of these unique war trauma injuries have not been described since the 1970s., Methods: Retrospective record review of all trauma casualties 5 to 65 years of age evacuated from the Iraqi theatre to U.S. Navy hospital ship, USNS Comfort, March to May 2003.War trauma-associated infection was defined by positive culture from a wound or sterile body fluid (ie, blood, cerebrospinal fluid) and at least two of the following infection-associated signs/symptoms: fever, dehiscence, foul smell, peri-wound erythema, hypotension, and leukocytosis. A comparison of mechanisms of injury, demographics, and clinical variables was done using multivariate analysis., Results: Of 211 patients, 56 met criteria for infection. Infections were more common in blast injuries, soft tissue injuries, >3 wound sites, loss of limb, abdominal trauma, and higher Injury Severity Score (ISS). Wound infections accounted for 84% of cases, followed by bloodstream infections (38%). Infected were more likely to have had fever prior to arrival, and had higher probability of ICU admission and more surgical procedures. Acinetobacter species (36%) were the predominant organisms followed by Escherichia coli and Pseudomonas species (14% each)., Conclusions: Similar to the Vietnam War experience, gram-negative rods, particularly Acinetobacter species, accounted for the majority of wound infections cared for on USNS Comfort during Operation Iraqi Freedom. Multidrug resistance was common, with the exception of the carbapenem class, limiting antibiotic therapy options.