A Role for NKG2D in NK Cell–Mediated Resistance to Poxvirus Disease

Ectromelia virus (ECTV) is an orthopoxvirus (OPV) that causes mousepox, the murine equivalent of human smallpox. C57BL/6 (B6) mice are naturally resistant to mousepox due to the concerted action of innate and adaptive immune responses. Previous studies have shown that natural killer (NK) cells are a component of innate immunity that is essential for the B6 mice resistance to mousepox. However, the mechanism of NK cell–mediated resistance to OPV disease remains undefined. Here we show that B6 mice resistance to mousepox requires the direct cytolytic function of NK cells, as well as their ability to boost the T cell response. Furthermore, we show that the activating receptor NKG2D is required for optimal NK cell–mediated resistance to disease and lethality. Together, our results have important implication towards the understanding of natural resistance to pathogenic viral infections

Influence of Soldiers' Cardiorespiratory Fitness on Physiological Responses and Dropouts During a Loaded Long-distance March.

Introduction: In military service, marching is an important, common, and physically demanding task. Minimizing dropouts, maintaining operational readiness during the march, and achieving a fast recovery are desirable because the soldiers have to be ready for duty, sometimes shortly after an exhausting task. The present field study investigated the influence of the soldiers' cardiorespiratory fitness on physiological responses during a long-lasting and challenging 34 km march. Materials and methods: Heart rate (HR), body core temperature (BCT), total energy expenditure (TEE), energy intake, motivation, and pain sensation were investigated in 44 soldiers (20.3 ± 1.3 years, 178.5 ± 7.0 cm, 74.8 ± 9.8 kg, body mass index: 23.4 ± 2.7 kg × m-2, peak oxygen uptake ($\dot{\rm{V}}$O2peak): 54.2 ± 7.9 mL × kg-1 × min-1) during almost 8 hours of marching. All soldiers were equipped with a portable electrocardiogram to record HR and an accelerometer on the hip, all swallowed a telemetry pill to record BCT, and all filled out a pre- and post-march questionnaire. The influence of aerobic capacity on the physiological responses during the march was examined by dividing the soldiers into three fitness groups according to their $\dot{\rm{V}}$O2peak. Results: The group with the lowest aerobic capacity ($\dot{\rm{V}}$O2peak: 44.9 ± 4.8 mL × kg-1 × min-1) compared to the group with the highest aerobic capacity ($\dot{\rm{V}}$O2peak: 61.7 ± 2.2 mL × kg-1 × min-1) showed a significantly higher (P < .05) mean HR (133 ± 9 bpm and 125 ± 8 bpm, respectively) as well as peak BCT (38.6 ± 0.3 and 38.4 ± 0.2 °C, respectively) during the march. In terms of recovery ability during the break, no significant differences could be identified between the three groups in either HR or BCT. The energy deficit during the march was remarkably high, as the soldiers could only replace 22%, 26%, and 36% of the total energy expenditure in the lower, middle, and higher fitness group, respectively. The cardiorespiratory fittest soldiers showed a significantly higher motivation to perform when compared to the least cardiorespiratory fit soldiers (P = .002; scale from 1 [not at all] to 10 [extremely]; scale difference of 2.3). A total of nine soldiers (16%) had to end marching early: four soldiers (21%) in the group with the lowest aerobic capacity, five (28%) in the middle group, and none in the highest group. Conclusion: Soldiers with a high $\dot{\rm{V}}$O2peak showed a lower mean HR and peak BCT throughout the long-distance march, as well as higher performance motivation, no dropouts, and lower energy deficit. All soldiers showed an enormous energy deficit; therefore, corresponding nutritional strategies are recommended

Conventional and regulatory CD4+ T cells that share identical TCRs are derived from common clones

Results from studies comparing the diversity and specificity of the TCR repertoires expressed by conventional (Tconv) and regulatory (Treg) CD4+ T cell have varied depending on the experimental system employed. We developed a new model in which T cells express a single fixed TCRα chain, randomly rearranged endogenous TCRβ chains, and a Foxp3-GFP reporter. We purified CD4+Foxp3- and CD4+Foxp3+ cells, then performed biased controlled multiplex PCR and high throughput sequencing of endogenous TCRβ chains. We identified >7,000 different TCRβ sequences in the periphery of 5 individual mice. On average, ~12% of TCR sequences were expressed by both conventional and regulatory populations within individual mice. The CD4+ T cells that expressed shared TCR sequences were present at higher frequencies compared to T cells expressing non-shared TCRs. Furthermore, nearly all (>90%) of the TCR sequences that were shared within mice were identical at the DNA sequence level, indicating that conventional and regulatory T cells that express shared TCRs are derived from common clones. Analysis of TCR repertoire overlap in the thymus reveals that a large proportion of Tconv and Treg sharing observed in the periphery is due to clonal expansion in the thymus. Together these data show that there are a limited number of TCR sequences shared between Tconv and Tregs. Also, Tconv and Tregs sharing identical TCRs are found at relatively high frequencies and are derived from common progenitors, of which a large portion are generated in the thymus

Poxvirus Bioinformatics Resource Center: a comprehensive Poxviridae informational and analytical resource

The Poxvirus Bioinformatics Resource Center (PBRC) has been established to provide informational and analytical resources to the scientific community to aid research directed at providing a better understanding of the Poxviridae family of viruses. The PBRC was specifically established as the result of the concern that variola virus, the causative agent of smallpox, as well as related viruses, might be utilized as biological weapons. In addition, the PBRC supports research on poxviruses that might be considered new and emerging infectious agents such as monkeypox virus. The PBRC consists of a relational database and web application that supports the data storage, annotation, analysis and information exchange goals of the project. The current release consists of over 35 complete genomic sequences of various genera, species and strains of viruses from the Poxviridae family. Sequence and annotation information for these viruses has been obtained from sequences publicly available from GenBank as well as sequences not yet deposited in GenBank that have been obtained from ongoing sequencing projects. In addition to sequence data, the PBRC provides comprehensive annotation and curation of virus genes; analytical tools to aid in the understanding of the available sequence data, including tools for the comparative analysis of different virus isolates; and visualization tools to help better display the results of various analyses. The PBRC represents the initial development of what will become a more comprehensive Viral Bioinformatics Resource Center for Biodefense that will be one of the National Institute of Allergy and Infectious Diseases' ‘Bioinformatics Resource Centers for Biodefense and Emerging or Re-Emerging Infectious Diseases’. The PBRC website is available at http://www.poxvirus.org

Validation of ambulatory monitoring devices to measure energy expenditure and heart rate in a military setting

Objectives.; To investigate the validity of different devices and algorithms used in military organizations worldwide to assess physical activity energy expenditure (PAEE) and heart rate (HR) among soldiers.; Design.; Device validation study.; Methods; . Twenty-three male participants serving their mandatory military service accomplished, firstly, nine different military specific activities indoors, and secondly, a normal military routine outdoors. Participants wore simultaneously an ActiHeart, Everion, MetaMax 3B, Garmin Fenix 3, Hidalgo EQ02, and PADIS 2.0 system. The PAEE and HR data of each system were compared to the criterion measures MetaMax 3B and Hidalgo EQ02, respectively.; Results; . Overall, the recorded systematic errors in PAEE estimation ranged from 0.1 (±1.8) kcal.min; -1; to -1.7 (±1.8) kcal.min; -1; for the systems PADIS 2.0 and Hidalgo EQ02 running the Royal Dutch Army algorithm, respectively, and in the HR assessment ranged from -0.1 (±2.1) b.min; -1; to 0.8 (±3.0) b.min; -1; for the PADIS 2.0 and ActiHeart systems, respectively. The mean absolute percentage error (MAPE) in PAEE estimation ranged from 29.9% to 75.1%, with only the Everion system showing an overall MAP

Whole-genome sequencing of a quarter-century melioidosis outbreak in temperate Australia uncovers a region of low-prevalence endemicity

This study was funded by the National Health and Medical Research Council via awards 1046812 and 1098337, and the Wellcome Trust Sanger Institute via award 098051. S.J.P. receives funding from the NIHR Cambridge Biomedical Research Centre.Melioidosis, caused by the highly recombinogenic bacterium Burkholderia pseudomallei, is a disease with high mortality. Tracing the origin of melioidosis outbreaks and understanding how the bacterium spreads and persists in the environment are essential to protecting public and veterinary health and reducing mortality associated with outbreaks. We used whole-genome sequencing to compare isolates from a historical quarter-century outbreak that occurred between 1966 and 1991 in the Avon Valley, Western Australia, a region far outside the known range of B. pseudomallei endemicity. All Avon Valley outbreak isolates shared the same multilocus sequence type (ST-284), which has not been identified outside this region. We found substantial genetic diversity among isolates based on a comparison of genome-wide variants, with no clear correlation between genotypes and temporal, geographical or source data. We observed little evidence of recombination in the outbreak strains, indicating that genetic diversity among these isolates has primarily accrued by mutation. Phylogenomic analysis demonstrated that the isolates confidently grouped within the Australian B. pseudomallei clade, thereby ruling out introduction from a melioidosis-endemic region outside Australia. Collectively, our results point to B. pseudomallei ST-284 being present in the Avon Valley for longer than previously recognized, with its persistence and genomic diversity suggesting long-term, low-prevalence endemicity in this temperate region. Our findings provide a concerning demonstration of the potential for environmental persistence of B. pseudomallei far outside the conventional endemic regions. An expected increase in extreme weather events may reactivate latent B. pseudomallei populations in this region.Publisher PDFPeer reviewe

Genomic sequence and analysis of a vaccinia virus isolate from a patient with a smallpox vaccine-related complication

BACKGROUND: Vaccinia virus (VACV)-DUKE was isolated from a lesion on a 54 year old female who presented to a doctor at the Duke University Medical Center. She was diagnosed with progressive vaccinia and treated with vaccinia immune globulin. The availability of the VACV-DUKE genome sequence permits a first time genomic comparison of a VACV isolate associated with a smallpox vaccine complication with the sequence of culture-derived clonal isolates of the Dryvax vaccine. RESULTS: This study showed that VACV-DUKE is most similar to VACV-ACAM2000 and CLONE3, two VACV clones isolated from the Dryvax(® )vaccine stock confirming VACV-DUKE as an isolate from Dryvax(®). However, VACV-DUKE is unique because it is, to date, the only Dryvax(® )clone isolated from a patient experiencing a vaccine-associated complication. The 199,960 bp VACV-DUKE genome encodes 225 open reading frames, including 178 intact genes and 47 gene fragments. Between VACV-DUKE and the other Dryvax(® )isolates, the major genomic differences are in fragmentation of the ankyrin-like, and kelch-like genes, presence of a full-length Interferon-α/β receptor gene, and the absence of a duplication of 12 ORFs in the inverted terminal repeat. Excluding this region, the DNA sequence of VACV-DUKE differs from the other two Dryvax(® )isolates by less than 0.4%. DNA sequencing also indicated that there was little heterogeneity in the sample, supporting the hypothesis that virus from an individual lesion is clonal in origin despite the fact that the vaccine is a mixed population. CONCLUSION: Virus in lesions that result from progressive vaccinia following vaccination with Dryvax are likely clonal in origin. The genomic sequence of VACV-DUKE is overall very similar to that of Dryvax(® )cell culture-derived clonal isolates. Furthermore, with the sequences of multiple clones from Dryvax(® )we can begin to appreciate the diversity of the viral population in the smallpox vaccine

High Efficiency Planar Geometry Germanium-on-silicon Single-photon Avalanche Diode Detectors

This paper presents the performance of 26 μm and 50 μm diameter planar Ge-on-Si single-photon avalanche diode (SPAD) detectors. The addition of germanium in these detectors extends the spectral range into the short-wave infrared (SWIR) region, beyond the capability of already well-established Si SPAD devices. There are several advantages for extending the spectral range into the SWIR region including: reduced eye-safety laser threshold, greater attainable ranges, and increased depth resolution in range finding applications, in addition to the enhanced capability to image through obscurants such as fog and smoke. The time correlated single-photon counting (TCSPC) technique has been utilized to observe record low dark count rates, below 100 kHz at a temperature of 125 K for up to a 6.6 % excess bias, for the 26 μm diameter devices. Under identical experimental conditions, in terms of excess bias and temperature, the 50 μm diameter device consistently demonstrates dark count rates a factor of 4 times greater than 26 μm diameter devices, indicating that the dark count rate is proportional to the device volume. Single-photon detection efficiencies of up to ~ 29 % were measured at a wavelength of 1310 nm at 125 K. Noise equivalent powers (NEP) down to 9.8 × 10-17 WHz-1/2 and jitters &lt; 160 ps are obtainable, both significantly lower than previous 100 μm diameter planar geometry devices, demonstrating the potential of these devices for highly sensitive and high-speed imaging in the SWIR

RNA helicase signaling is critical for type I interferon production and protection against rift valley fever virus during mucosal challenge

Rift Valley fever virus (RVFV) is an emerging RNA virus with devastating economic and social consequences. Clinically, RVFV induces a gamut of symptoms ranging from febrile illness to retinitis, hepatic necrosis, hemorrhagic fever, and death. It is known that type I interferon (IFN) responses can be protective against severe pathology; however, it is unknown which innate immune receptor pathways are crucial for mounting this response. Using both in vitro assays and in vivo mucosal mouse challenge, we demonstrate here that RNA helicases are critical for IFN production by immune cells and that signaling through the helicase adaptor molecule MAVS (mitochondrial antiviral signaling) is protective against mortality and more subtle pathology during RVFV infection. In addition, we demonstrate that Toll-like-receptor-mediated signaling is not involved in IFN production, further emphasizing the importance of the RNA cellular helicases in type I IFN responses to RVFV