Hantavirus Pulmonary Syndrome: CD8+and CD4+Cytotoxic T Lymphocytes to Epitopes on Sin Nombre Virus Nucleocapsid Protein Isolated during Acute Illness

AbstractIn 1993 a number of cases of unexplained adult respiratory syndrome occurred in the southwestern United States. The illness was characterized by a prodrome of fever, myalgia, and other symptoms followed by the rapid onset of a capillary leak syndrome with hemoconcentration, thrombocytopenia, and pulmonary edema. Viral RNA sequences in the lungs identified a new member of the hantavirus genus, Sin Nombre virus (SNV), unique to North America. Pulmonary endothelial cells were heavily infected but were not necrotic. We speculated that this capillary leak syndrome was initiated by immune responses to the SNV-infected pulmonary endothelial cells. We isolated a CD8+cytotoxic T lymphocyte (CTL) clone directly from the blood of a patient with the acute hantavirus pulmonary syndrome (HPS) which recognizes a SNV specific epitope on the virus nucleocapsid protein (aa 234–242) that is restricted by HLA C7 and produces IFNγ but not IL-4. We identified a second CD8+CTL epitope located within another site aa 131–139 on the nucleocapsid protein, which is HLA B35 restricted, and a CD4+CTL epitope located on a third site on nucleocapsid protein aa 372–380 using lymphocytes obtained during HPS from another patient that were stimulatedin vitro.Hantavirus specific CD8+and CD4+CTL may contribute to the immunopathology and capillary leak syndrome observed in the HPS

Transmission of Aerosolized Seasonal H1N1 Influenza A to Ferrets

Influenza virus is a major cause of morbidity and mortality worldwide, yet little quantitative understanding of transmission is available to guide evidence-based public health practice. Recent studies of influenza non-contact transmission between ferrets and guinea pigs have provided insights into the relative transmission efficiencies of pandemic and seasonal strains, but the infecting dose and subsequent contagion has not been quantified for most strains. In order to measure the aerosol infectious dose for 50% (aID50) of seronegative ferrets, seasonal influenza virus was nebulized into an exposure chamber with controlled airflow limiting inhalation to airborne particles less than 5 µm diameter. Airborne virus was collected by liquid impinger and Teflon filters during nebulization of varying doses of aerosolized virus. Since culturable virus was accurately captured on filters only up to 20 minutes, airborne viral RNA collected during 1-hour exposures was quantified by two assays, a high-throughput RT-PCR/mass spectrometry assay detecting 6 genome segments (Ibis T5000™ Biosensor system) and a standard real time RT-qPCR assay. Using the more sensitive T5000 assay, the aID50 for A/New Caledonia/20/99 (H1N1) was approximately 4 infectious virus particles under the exposure conditions used. Although seroconversion and sustained levels of viral RNA in upper airway secretions suggested established mucosal infection, viral cultures were almost always negative. Thus after inhalation, this seasonal H1N1 virus may replicate less efficiently than H3N2 virus after mucosal deposition and exhibit less contagion after aerosol exposure

Enhanced Immunogenicity, Mortality Protection, and Reduced Viral Brain Invasion by Alum Adjuvant with an H5N1 Split-Virion Vaccine in the Ferret

Pre-pandemic development of an inactivated, split-virion avian influenza vaccine is challenged by the lack of pre-existing immunity and the reduced immunogenicity of some H5 hemagglutinins compared to that of seasonal influenza vaccines. Identification of an acceptable effective adjuvant is needed to improve immunogenicity of a split-virion avian influenza vaccine.No serum antibodies were detected after vaccination with unadjuvanted vaccine, whereas alum-adjuvanted vaccination induced a robust antibody response. Survival after unadjuvanted dose regimens of 30 µg, 7.5 µg and 1.9 µg (21-day intervals) was 64%, 43%, and 43%, respectively, yet survivors experienced weight loss, fever and thrombocytopenia. Survival after unadjuvanted dose regimen of 22.5 µg (28-day intervals) was 0%, suggesting important differences in intervals in this model. In contrast to unadjuvanted survivors, either dose of alum-adjuvanted vaccine resulted in 93% survival with minimal morbidity and without fever or weight loss. The rarity of brain inflammation in alum-adjuvanted survivors, compared to high levels in unadjuvanted vaccine survivors, suggested that improved protection associated with the alum adjuvant was due to markedly reduced early viral invasion of the ferret brain.Alum adjuvant significantly improves efficacy of an H5N1 split-virion vaccine in the ferret model as measured by immunogenicity, mortality, morbidity, and brain invasion

Exhaled Aerosol Transmission of Pandemic and Seasonal H1N1 Influenza Viruses in the Ferret

Person-to-person transmission of influenza viruses occurs by contact (direct and fomites) and non-contact (droplet and small particle aerosol) routes, but the quantitative dynamics and relative contributions of these routes are incompletely understood. The transmissibility of influenza strains estimated from secondary attack rates in closed human populations is confounded by large variations in population susceptibilities. An experimental method to phenotype strains for transmissibility in an animal model could provide relative efficiencies of transmission. We developed an experimental method to detect exhaled viral aerosol transmission between unanesthetized infected and susceptible ferrets, measured aerosol particle size and number, and quantified the viral genomic RNA in the exhaled aerosol. During brief 3-hour exposures to exhaled viral aerosols in airflow-controlled chambers, three strains of pandemic 2009 H1N1 strains were frequently transmitted to susceptible ferrets. In contrast one seasonal H1N1 strain was not transmitted in spite of higher levels of viral RNA in the exhaled aerosol. Among three pandemic strains, the two strains causing weight loss and illness in the intranasally infected ‘donor’ ferrets were transmitted less efficiently from the donor than the strain causing no detectable illness, suggesting that the mucosal inflammatory response may attenuate viable exhaled virus. Although exhaled viral RNA remained constant, transmission efficiency diminished from day 1 to day 5 after donor infection. Thus, aerosol transmission between ferrets may be dependent on at least four characteristics of virus-host relationships including the level of exhaled virus, infectious particle size, mucosal inflammation, and viral replication efficiency in susceptible mucosa

Genetic Determinants of Serum Testosterone Concentrations in Men

Testosterone concentrations in men are associated with cardiovascular morbidity, osteoporosis, and mortality and are affected by age, smoking, and obesity. Because of serum testosterone's high heritability, we performed a meta-analysis of genome-wide association data in 8,938 men from seven cohorts and followed up the genome-wide significant findings in one in silico (n = 871) and two de novo replication cohorts (n = 4,620) to identify genetic loci significantly associated with serum testosterone concentration in men. All these loci were also associated with low serum testosterone concentration defined as <300 ng/dl. Two single-nucleotide polymorphisms at the sex hormone-binding globulin (SHBG) locus (17p13-p12) were identified as independently associated with serum testosterone concentration (rs12150660, p = 1.2×10−41 and rs6258, p = 2.3×10−22). Subjects with ≥3 risk alleles of these variants had 6.5-fold higher risk of having low serum testosterone than subjects with no risk allele. The rs5934505 polymorphism near FAM9B on the X chromosome was also associated with testosterone concentrations (p = 5.6×10−16). The rs6258 polymorphism in exon 4 of SHBG affected SHBG's affinity for binding testosterone and the measured free testosterone fraction (p<0.01). Genetic variants in the SHBG locus and on the X chromosome are associated with a substantial variation in testosterone concentrations and increased risk of low testosterone. rs6258 is the first reported SHBG polymorphism, which affects testosterone binding to SHBG and the free testosterone fraction and could therefore influence the calculation of free testosterone using law-of-mass-action equation

Food and Nutrition Board National Research Council

Reporte de investigación -- Universidad de Costa Rica, Instituto de Investigaciones en Salud. 1978The magnitude of malnutrition in the world is staggering. In addition to its major causative factors, which relate specifically to food availability at the household level, infectious diseases are an omnipresent and important inciter and contributor to malnutrition and its associated morbidity and mortality. Therefore, there are many factors that deserve attention during any examination of the malnutrition-infection complex. This report addresses only those factors expected to reduce the impact and burden of infectious diseases as a means to improving nutritional status and concentrates on recommendations for field-level implementation programs. To accomplish this, the subcommittee examined the following: • the interaction of nutrition and infection, which is called the malnutrition-infection complex; • targeting of programs to specific subsegments of the population, specifically to pregnant and lactating women, weanlings and children less than three years of age; • incorporation of adaptive research and impact evaluation into the program design; and • national planning and policymaking, local-level responsibility, outreach to the household level, and the establishment of linkages between the national and local activities . The subcommittee recommends that four elements be included in the program as a "package": • Prenatal control to increase maternal caloric intake through education and/or supplements (preferably using indigenous foods), however provided; to eradicate neonatal tetanus through immunization of the mother; to treat nutritional anemia and other specific deficiencies;to monitor weight gain by distributing take-home charts on which infant birth weight and growth can be recorded; and to initiate nutrition education, including concepts of breast-feeding, maternal requirements for lactation, weaning foods, oral rehydration therapy for dehydration, and postinfection convalescent nutritional care. • Emphasis on promoting breast-feeding, prolonging the practice as long as possible, and introducingsupplemental feeding practices with locally prepared indigenous foods. Completed growth charts and calculation of incremental growth will assist in the identification of growth-faltering infants, a group requiring extra attention. • Introduction of oral rehydration into households and instruction in its proper use in early therapy for acute diarrhea, along with continuation or early reinstitution of feeding for nutritional maintenance. • Programs to immunize children less than one-year-old with DPT (diphtheria-pertussis-tetanus), BeG (bacille Calmette Guerin), measles, and polio vaccines while the child is adequately nourished (as established by growth surveillance) and to maintain immunization records by means of growth charts that are completed in the home. Because there should be no dichotomy between doing and learning, the subcommittee recommends that the program include a research arm that would seek the information necessary to shape and define efforts to achieve maximum efficiency and success.Universidad de Costa Rica. National Academy of Sciences. This study was supported by Contract AID/TA-C-1428 with the Agency for International Development.UCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto de Investigaciones en Salud (INISA

Role of specific CD8+ T cells in the severity of a fulminant zoonotic viral hemorrhagic fever, hantavirus pulmonary syndrome

We report on the role of specific CD8(+) T cells in the pathogenesis of a highly lethal human viral disease, hantavirus pulmonary syndrome (HPS). HPS is a zoonotic disease caused by transmission of Sin Nombre virus (SNV) from chronically infected deer mice. In humans, this fulminant infection is characterized by lung capillary leakage, respiratory failure, and cardiogenic shock. Individuals with HLA-B*3501 have an increased risk of developing severe HPS, suggesting that CD8(+) T cell responses to SNV contribute to pathogenesis. We identified three CD8(+) T cell epitopes in SNV presented by HLA-B*3501 and quantitated circulating SNV-specific CD8(+) T cells in 11 acute HPS patients using HLA/peptide tetramers. We found significantly higher frequencies of SNV-specific T cells in patients with severe HPS requiring mechanical ventilation (up to 44.2% of CD8(+) T cells) than in moderately ill HPS patients hospitalized but not requiring mechanical ventilation (up to 9.8% of CD8(+) T cells). These results imply that virus-specific CD8(+) T cells contribute to HPS disease outcome. Intense CD8(+) T cell responses to SNV may be induced by the encounter of the unnatural human host to this zoonotic virus without coevolution. This may also be the immunopathologic basis of other life-threatening human virus infections