Saliva Preservative for Diagnostic Purposes

Saliva is an important body fluid for diagnostic purposes. Glycoproteins, glucose, steroids, DNA, and other molecules of diagnostic value are found in saliva. It is easier to collect as compared to blood or urine. Unfortunately, saliva also contains large numbers of bacteria that can release enzymes, which can degrade proteins and nucleic acids. These degradative enzymes destroy or reduce saliva s diagnostic value. This innovation describes the formulation of a chemical preservative that prevents microbial growth and inactivates the degradative enzymes. This extends the time that saliva can be stored or transported without losing its diagnostic value. Multiple samples of saliva can be collected if needed without causing discomfort to the subject and it does not require any special facilities to handle after it is collected

Latent Herpes Viruses Reactivation in Astronauts

Space flight has many adverse effects on human physiology. Changes in multiple systems, including the cardiovascular, musculoskeletal, neurovestibular, endocrine, and immune systems have occurred (12, 32, 38, 39). Alterations in drug pharmacokinetics and pharmacodynamics (12), nutritional needs (31), renal stone formation (40), and microbial flora (2) have also been reported. Evidence suggests that the magnitude of some changes may increase with time in space. A variety of changes in immunity have been reported during both short (.16 days) and long (>30 days) space missions. However, it is difficult to determine the medical significance of these immunological changes in astronauts. Astronauts are in excellent health and in superb physical condition. Illnesses in astronauts during space flight are not common, are generally mild, and rarely affect mission objectives. In an attempt to clarify this issue, we identified the latent herpes viruses as medically important indicators of the effects of space flight on immunity. This chapter demonstrates that space flight leads to asymptomatic reactivation of latent herpes viruses, and proposes that this results from marked changes in neuroendocrine function and immunity caused by the inherent stressfullness of human space flight. Astronauts experience uniquely stressful environments during space flight. Potential stressors include confinement in an unfamiliar, crowded environment, isolation, separation from family, anxiety, fear, sleep deprivation, psychosocial issues, physical exertion, noise, variable acceleration forces, increased radiation, and others. Many of these are intermittent and variable in duration and intensity, but variable gravity forces (including transitions from launch acceleration to microgravity and from microgravity to planetary gravity) and variable radiation levels are part of each mission and contribute to a stressful environment that cannot be duplicated on Earth. Radiation outside the Earth's magnetosphere is particularly worrisome because it includes ionizing radiation from cosmic galactic radiation. Increased stress levels appear even before flight, presumably from the rigors of preflight training and the anticipation of the mission (12, 32, 38, 39). Space flight causes significant changes in human immune function (32), but the means by which these changes come about have been difficult to discern. Consistent indicators of stress associated with space flight include increased production of stress hormones, and changes in cells of the immune system. These changes include elevated white blood cell (WBC) and neutrophil counts at landing (15, 16, 35, 37). Activation of generalized stress responses before, during, and after space flight probably affects the function of the immune system. Space flight has been shown to decrease many aspects of immune function, including natural killer (NK) cell activity, interferon production, the blastogenic response of leukocytes to mitogens, cell-mediated immunity, neutrophil function and monocyte function (5, 16, 18, 21, 35-37)

Spaceflight modulates gene expression in the whole blood of astronauts

Astronauts are exposed to a unique combination of stressors during spaceflight, which leads to alterations in their physiology and potentially increases their susceptibility to disease, including infectious diseases. To evaluate the potential impact of the spaceflight environment on the regulation of molecular pathways mediating cellular stress responses, we performed a first-of-its-kind pilot study to assess spaceflight-related gene-expression changes in the whole blood of astronauts. Using an array comprised of 234 well-characterized stress-response genes, we profiled transcriptomic changes in six astronauts (four men and two women) from blood preserved before and immediately following the spaceflight. Differentially regulated transcripts included those important for DNA repair, oxidative stress, and protein folding/degradation, including HSP90AB1, HSP27, GPX1, XRCC1, BAG-1, HHR23A, FAP48, and C-FOS. No gender-specific differences or relationship to number of missions flown was observed. This study provides a first assessment of transcriptomic changes occurring in the whole blood of astronauts in response to spaceflight

Determination of Roles of Microgravity and Ionizing Radiation on the Reactivation of Epstein-Barr Virus In Vitro

Astronauts experience symptomatic and asymptomatic herpes virus reactivation during spaceflight. We have shown increases in reactivation of Epstein-Barr virus (EBV), cytomegalovirus (CMV) and varicella zoster virus (VZV) and shedding in body fluids (saliva and urine) in astronauts during space travel. Alterations in immunity, increased stress hormone levels, microgravity, increased radiation, and other conditions unique to spaceflight may promote reactivation of latent herpes viruses. Unique mechanico-physico forces associated with spaceflight can have profound effects on cellular function, especially immune cells. In space flight analog studies such as Antarctica, bed rest studies, and NASA's undersea habitat (Aquarius), reactivation of these viruses occurred, but to a lesser extent than spaceflight. Spaceflight analogs model some spaceflight factors, but none of the analogs recreates all factors experienced in space. Most notably, microgravity and radiation are not included in many analogs. Stress, processed through the HPA axis and SAM systems, induces viral reactivation. However, the respective roles of microgravity and increased space radiation levels or if any synergy exists are not known. Therefore, we studied the effect of modeled space radiation and/or microgravity, independent of the immune system on the changes in cellular gene expression that results in viral (EBV) reactivation. The effects of modeled microgravity and low shear on EBV replication and cellular and EBV gene expression were studied in human B-lymphocyte cell cultures. Latently infected B-lymphocytes were propagated in the rotating wall bioreactor and irradiated with the various dosages of gamma irradiation. At specific time intervals following exposure to modeled microgravity, the cells and supernatant were harvested and reactivation of EBV were assessed by measuring EBV and gene expression, DNA methylation, and infectious virus production

Genetic Diversity Analysis of Mutant Lines of Oat (\u3cem\u3eAvena sativa\u3c/em\u3e L.) Based on RAPD and ISSR Analysis

The genus Avena belongs to the grass family Poaceae and has ploidy levels of diploid, tetraploid and hexaploid with basic chromosome number of 7 (n=7). Oat (Avena sativa L.) is one of the most important forage and feed crops of the world. Oat is used as green fodder, straw, hay or silage. Oat grain makes a good balanced concentrate in the rations for poultry, cattle, sheep and other animals. Green fodder contains about 10 to 13% protein and 30 to 35% dry matter. Despite being high fed fodder crop, it is now gaining importance as food due to its unique and important quality characteristics, particularly the lipid and protein in grains (Ruwali et al., 2013). The existing genetic variability for the traits of agronomic importance, such as plant vegetative cycle, is considered restricted. The narrow of the genetic base in cultivated oat varieties can be a constraint on the efficacy of genotype selection in segregating generations (Carvalho and Federizzi, 1989). Genetic variability in existing oat cultivars is not high enough; it hampers the selection of superior genotypes for breeding. Modifications in the genetic structure of plants and an organisms occurs naturally, though at low frequency, but can be increased through physical or chemical mutagens. Advances in molecular biology have introduced an alternative for variety/genotype identification. The genetic characterization of germplasm helps in their effective conservation and reveals the extent of relationship among the accessions and the estimates of genetic diversity (Singh et al., 2012). The selection of RAPD and ISSR were based on their relative technical simplicity, level of polymorphism they detect, cost effective, easily applicable to any plant species and target those sequence which are abundant throughout the eukaryotic genome and are rapidly evolved. A series of studies have indicated that ISSR could be able to produce more reliable and reproducible bands because of the higher annealing temperature and longer sequence of ISSR primers considered superior than RAPD (Bornet et al., 2001). ISSR has proved to be useful to study of population genetic studies gene mapping germplasm identification and characterize gene bank accessions as well as to identify closely related cultivars (Cortesi et al., 2004). The present research had the following objectives: Assessment of diversity of mutant lines of oat (Avena sativa L.) based on RAPD and ISSR analysis

Varicella-Zoster DNA in Saliva of Patients with Meningoencephalitis - A Preliminary Study

No abstract availabl

The Impact of Long Duration Spaceflight on Plasma Antimicrobial Proteins

Introduction: Robust immunity is essential for further human exploration of the solar system beyond Earth’s orbit. Spaceflight has been associated with immune perturbations and latent viral reactivation. However, logistical constraints have restricted many of these studies to simple pre- and post-flight measures, which are greatly confounded by the stressors associated with launch, landing and re-adaptation to the 1G environment. More in-flight immune data are required particularly during long-duration (3-6 months) spaceflight missions. This study examined the effects of spaceflight on plasma antimicrobial proteins (AMPs) and reactivation of latent herpesviruses. Methods: Plasma, saliva and urine samples were obtained from 20 crewmembers who spent ~6-months on the International Space Station (ISS). Samples were collected 180 and 45-days before launch, in-flight (at ‘early, ‘mid’ and ‘late’ stages of the mission), immediately upon return to Earth (R+0) and 30 days following return (R+30). Plasma LL-37, HNP 1-3 and lysozyme concentrations were determined by ELISA. Saliva Epstein-Barr virus (EBV), varicella zoster virus (VZV) and urine cytomegalovirus (CMV) DNA levels were quantified by Real-Time PCR. Maximum likelihood linear mixed models (LMM) were used to determine main effects of time (pre-flight, in-flight, R+0 and R+30), and EBV, VZV and CMV viral shedding status (shedding or non-shedding) on the concentration of each AMPs. Results: Lower plasma levels of LL-37 were found at R+0, compared to pre-flight, in-flight and R+30 (-80.6%, -80.2% and -73.49% respectively; p \u3c 0.01). Plasma HNP 1-3 levels were elevated above pre-flight level during flight, at R+0 and R+30 (+24%, +40% and +17% respectively; p \u3c 0.01). Only those crewmembers found to shed CMV had a significant reduction in plasma LL-37 at R+0 (p \u3c 0.05). Similarly, crewmembers found to shed VZV at R+0 had lower HNP 1-3 concentrations than crewmembers who did not shed VZV (-68.9%; p \u3c 0.01). Finally, only those crewmembers who shed EBV had increased plasma levels of HNP 1-3 at R+0 (p \u3c 0.01). Plasma lysozyme levels were unaffected by spaceflight or latent viral shedding. Conclusion: Long-duration spaceflight alters plasma LL-37 and HNP 1-3 levels and are linked to the reactivation of latent herpesviruses. The in-flight changes observed for HNP 1-3 indicate that certain immune perturbations may be independent of launch/landing stress. Future studies are required to determine if spaceflight induced immune dysregulation increases the risk of an adverse health event before exploration-class planetary missions (i.e. to Mars) can be considered

Rapid Detection of the Varicella Zoster Virus in Saliva

Varicella zoster virus (VZV) causes chicken pox on first exposure (usually in children), and reactivates from latency causing shingles (usually in adults). Shingles can be extremely painful, causing nerve damage, organ damage, and blindness in some cases. The virus can be life-threatening in immune-compromised individuals. The virus is very difficult to culture for diagnosis, requiring a week or longer. This invention is a rapid test for VZV from a saliva sample and can be performed in a doctor s office. The kit is small, compact, and lightweight. Detec tion is sensitive, specific, and noninvasive (no needles); only a saliva sample is required. The test provides results in minutes. The entire test is performed in a closed system, with no exposure to infectious materials. The components are made mostly of inexpensive plastic injection molded parts, many of which can be purchased off the shelf and merely assembled. All biological waste is contained for fast, efficient disposal. This innovation was made possible because of discovery of a NASA scientists flight experiment showing the presence of VZV in saliva during high stress periods and disease. This finding enables clinicians to quickly screen patients for VZV and treat the ones that show positive results with antiviral medicines. This promotes a rapid recovery, easing of pain and symptoms, and reduces chances of complications from zoster. Screening of high-risk patients could be incorporated as part of a regular physical exam. These patients include the elderly, pregnant women, and immune-compromised individuals. In these patients, VZV can be a life-threatening disease. In both high- and low-risk patients, early detection and treatment with antiviral drugs can dramatically decrease or even eliminate the clinical manifestation of disease

Subclinical Shed of Infectious Varicella zoster Virus in Astronauts

Aerosol borne varicella zoster virus (VZV) enters the nasopharynx and replicates in tonsillar T-cells, resulting in viremia and varicella (chickenpox). Virus then becomes latent in cranial nerve, dorsal root and autonomic nervous system ganglia along the entire neuraxis (1). Decades later, as cell-mediated immunity to VZV declines (4), latent VZV can reactivate to produce zoster (shingles). Infectious VZV is present in patients with varicella or zoster, but shed of infectious virus in the absence of disease has not been shown. We previously detected VZV DNA in saliva of astronauts during and shortly after spaceflight, suggesting stress induced subclinical virus reactivation (3). We show here that VZV DNA as well as infectious virus in present in astronaut saliva. VZV DNA was detected in saliva during and after a 13-day spaceflight in 2 of 3 astronauts (Fig. panel A). Ten days before liftoff, there was a rise in serum anti-VZV antibody in subjects 1 and 2, consistent with virus reactivation. In subject 3, VZV DNA was not detected in saliva, and there was no rise in anti-VZV antibody titer. Subject 3 may have been protected from virus reactivation by having zoster <10 years ago, which provides a boost in cell-medicated immunity to VZV (2). No VZV DNA was detected in astronaut saliva months before spaceflight, or in saliva of 10 age/sex-matched healthy control subjects sampled on alternate days for 3 weeks (88 saliva samples). Saliva taken 2-6 days after landing from all 3 subjects was cultured on human fetal lung cells (Fig. panel B). Infectious VZV was recovered from saliva of subjects 1 and 2 on the second day after landing. Virus specificity was confirmed by antibody staining and DNA analysis which showed it to be VZV of European descent, common in the US (5). Further, both antibody staining and DNA PCR demonstrated that no HSV-1 was detected in any infected culture. This is the first report of infectious VZV shedding in the absence of clinical disease. Spaceflight presents a uniquely stressful environment which includes physical isolation and confinement, anxiety, sleep deprivation, as well as exposure to increased radiation and microgravity. It is interesting that in our study, VZV and not HSV-1 reactivation was detected, since stress-induced HSV-1 reactivation has been reported (6). Future studies are needed to determine the specific inducer of VZV reactivation

Stress-Induced Subclinical Reactivation of Varicella Zoster Virus in Astronauts

After primary infection, varicella-zoster virus (VZV) becomes latent in ganglia. VZV reactivation occurs primarily in elderly individuals, organ transplant recipients, and patients with cancer and AIDS, correlating with a specific decline in cell-mediated immunity to VZV. VZV can also reactivate after surgical stress. To determine whether VZV can also reactivate after acute non-surgical stress, we examined total DNA extracted from 312 saliva samples of eight astronauts before, during and after space flight for VZV DNA by PCR: 112 samples were obtained 234 to 265 days before flight, 84 samples on days 2 through 13 of space flight, and 116 samples on days 1 through 15 after flight. Before space flight only one of the 112 saliva samples from a single astronaut was positive for VZV DNA. In contrast, during and after space flight, 61 of 200 (30%) saliva samples were positive in all 8 astronauts. No VZV DNA was detected in any of 88 saliva samples from 10 healthy control subjects. These data indicate that VZV can reactivate subclinically in healthy individuals after acute stress