•A rapid sensitive method to obtain DNA from saliva allowed detection of VZV DNA in 100% of patients with acute herpes zoster before treatment.•Passive drool and synthetic swab yielded abundant ...amounts of total DNA.•High-speed centrifugation significantly reduced the time to process saliva from collection to obtaining DNA.•VZV DNA was found exclusively in the pelleted fraction of saliva.
VZV reactivation produces zoster (shingles) which may be further complicated by meningoencephalitis, myelopathy, vasculopathy and multiple ocular disorders. Importantly, these neurological and ocular complications of VZV reactivation can occur without rash. In such instances, virological verification relies on detection of VZV DNA or anti-VZV IgG antibody in cerebrospinal fluid (CSF), or less often, the presence of VZV DNA in blood mononuclear cells or anti-VZV IgM antibody in serum or CSF. If VZV were readily detected in other tissue samples (e.g., saliva or tears) in patients with neurological disease in the absence of rash and shown to correlate with the standard tests listed above, more invasive tests such as lumbar puncture might be obviated.
In patients with acute herpes zoster, the yield of cell DNA was greater in saliva collected by passive drool or synthetic swab than by cotton swab. The time to process saliva from collection to obtaining DNA was 1h. VZV DNA was present exclusively in the pelleted fraction of saliva and was found in 100% of patients before antiviral treatment. This rapid sensitive method can be applied readily to saliva from humans with neurologic and other disease that might be caused by VZV in the absence of rash.
Analysis of 36 individuals over age 60 years who were immunized with Zostavax revealed varicella zoster virus (VZV) DNA in swabs of skin inoculation sites obtained immediately after immunization in ...18 (50%) of 36 subjects (copy number per nanogram of total DNA, 28 to 2.1 × 10 6⁶) and in saliva collected over 28 days in 21 (58%) of 36 subjects (copy number, 20 to 248). Genotypic analysis of DNA extracted from 9 random saliva samples identified vaccine virus in all instances. In some immunized individuals over age 60, vaccine virus DNA is shed in saliva up to 4 weeks.
Most of our knowledge about how antiretrovirals and host immune responses influence the HIV-1 protease gene is derived from studies of subtype B virus. We investigated the effect of protease ...resistance-associated mutations (PRAMs) and population-based HLA haplotype frequencies on polymorphisms found in CRF01_AE pro.
We used all CRF01_AE protease sequences retrieved from the LANL database and obtained regional HLA frequencies from the dbMHC database. Polymorphisms and major PRAMs in the sequences were identified using the Stanford Resistance Database, and we performed phylogenetic and selection analyses using HyPhy. HLA binding affinities were estimated using the Immune Epitope Database and Analysis.
Overall, 99% of CRF01_AE sequences had at least 1 polymorphism and 10% had at least 1 major PRAM. Three polymorphisms (L10 V, K20RMI and I62 V) were associated with the presence of a major PRAM (P < 0.05). Compared to the subtype B consensus, six additional polymorphisms (I13 V, E35D, M36I, R41K, H69K, L89M) were identified in the CRF01_AE consensus; all but L89M were located within epitopes recognized by HLA class I alleles. Of the predominant HLA haplotypes in the Asian regions of CRF01_AE origin, 80% were positively associated with the observed polymorphisms, and estimated HLA binding affinity was estimated to decrease 19-40 fold with the observed polymorphisms at positions 35, 36 and 41.
Polymorphisms in CRF01_AE protease gene were common, and polymorphisms at residues 10, 20 and 62 most likely represent selection by use of protease inhibitors, whereas R41K and H69K were more likely attributable to recognition of epitopes by the HLA haplotypes of the host population.
Data on the detailed clinical progression of COVID-19 in conjunction with epidemiological and virological characteristics are limited. In this case series, we describe the first 12 US patients ...confirmed to have COVID-19 from 20 January to 5 February 2020, including 4 patients described previously
. Respiratory, stool, serum and urine specimens were submitted for SARS-CoV-2 real-time reverse-transcription polymerase chain reaction (rRT-PCR) testing, viral culture and whole genome sequencing. Median age was 53 years (range: 21-68); 8 patients were male. Common symptoms at illness onset were cough (n = 8) and fever (n = 7). Patients had mild to moderately severe illness; seven were hospitalized and demonstrated clinical or laboratory signs of worsening during the second week of illness. No patients required mechanical ventilation and all recovered. All had SARS-CoV-2 RNA detected in respiratory specimens, typically for 2-3 weeks after illness onset. Lowest real-time PCR with reverse transcription cycle threshold values in the upper respiratory tract were often detected in the first week and SARS-CoV-2 was cultured from early respiratory specimens. These data provide insight into the natural history of SARS-CoV-2. Although infectiousness is unclear, highest viral RNA levels were identified in the first week of illness. Clinicians should anticipate that some patients may worsen in the second week of illness.
Changes in leukocyte subpopulations and function after spaceflight have been observed but the mechanisms underlying these changes are not well defined. This study investigated the effects of ...short-term spaceflight (8-15 days) on circulating leukocyte subsets, stress hormones, immunoglobulin levels, and neutrophil function. At landing, a 1.5-fold increase in neutrophils was observed compared with preflight values; lymphocytes were slightly decreased, whereas the results were variable for monocytes. No significant changes were observed in plasma levels of immunoglobulins, cortisol, or adrenocorticotropic hormone. In contrast, urinary epinephrine, norepinephrine, and cortisol were significantly elevated at landing. Band neutrophils were observed in 9 of 16 astronauts. Neutrophil chemotactic assays showed a 10-fold decrease in the optimal dose response after landing. Neutrophil adhesion to endothelial cells was increased both before and after spaceflight. At landing, the expression of MAC-1 was significantly decreased while L-selectin was significantly increased. These functional alterations may be of clinical significance on long-duration space missions.
This pilot study compared placental growth factor (PIGF) levels in populations with high versus low risk for cardiovascular disease. Previous experiments from our laboratory (Sundaresan et al.
2005
,
...2009
) revealed that the angiogenic factor PIGF was up regulated in modeled microgravity conditions in human lymphocytes leading to possible atherogenesis and pathogenesis in microgravity. Since the findings came from microgravity analog experiments, there is a strong link to its usefulness in the microgravity field as a biomarker. It is important to understand, that these findings came from both studies on expression levels of this cardiovascular marker in human lymphocytes in microgravity (
in vitro
microgravity analog), and a follow up gene expression study in hind limb suspended mice (
in vivo
microgravity analog). The relevance is enhanced because in life on earth, PIGF is an inflammatory biomarker for cardiovascular disease. Studies on the levels of PIGF would help to reduce the risk and prevention of heart failures in astronauts. If we can use this marker to predict and reduce the risk of cardiac events in astronauts and pilots, it would significantly help aerospace medicine operations. The investigations here confirmed that in a cardiovascular stressed population such as coronary artery disease (CAD) and acute coronary syndrome (ACS) patients, PIGF could be overexpressed. We desired to re-evaluate this marker in patients with cardiovascular disease in our own study. PIGF is a marker of inflammation and a predictor of short-term and long-term adverse outcome in ACS. In addition, elevated PIGF levels may be associated with increased risk for CAD.PIGF levels were determined in thirty-one patients undergoing cardiovascular catheterization for reasons other than ACS and in thirty-three low-risk asymptomatic subjects. Additional data on traditional cardiovascular risk factors for both populations were also compiled and compared. We found that PIGF levels were significantly higher in the high-risk population as compared to low-risk population. Also we were able to ascertain that PIGF levels were inversely correlated with HDL-cholesterol but directly correlated with the triglyceride levels. With further validation, PIGF may prove a useful addition to the armamentarium of noninvasive biomarkers for cardiovascular disease including a new area of stressful physiological conditions such as microgravity.
Changes of gene expression profile are one of the most important biological responses in living cells after ionizing radiation (IR) exposure. Although some studies have shown that genes up-regulated ...by IR may play important roles in DNA damage repair, the relationship between the regulation of gene expression by IR, particularly genes not known for their roles in double-strand break (DSB) repair, and its impact on cytogenetic responses has not been well studied. The purpose of this study is to identify new roles of IR inducible genes in regulating DSB repair and cell cycle progression. In this study, the expression of 25 genes selected on the basis of their transcriptional changes in response to IR was individually knocked down by small interfering RNA in human fibroblast cells. Frequency of micronuclei (MN) formation and chromosome aberrations were measured to determine efficiency of cytogenetic repair, especially DSB repair. In response to IR, the formation of MN was significantly increased by suppressed expression of five genes: Ku70 (DSB repair pathway), XPA (nucleotide excision repair pathway), RPA1 (mismatch repair pathway), RAD17 and RBBP8 (cell cycle control). Knocked-down expression of four genes (MRE11A, RAD51 in the DSB pathway, SESN1, and SUMO1) significantly inhibited cell cycle progression, possibly because of severe impairment of DNA damage repair. Moreover, decreased XPA, p21, or MLH1 expression resulted in both significantly enhanced cell cycle progression and increased yields of chromosome aberrations, indicating that these gene products modulate both cell cycle control and DNA damage repair. Nine of these eleven genes, whose knock-down expression affected cytogenetic repair, were up-regulated in cells exposed to gamma radiation, suggesting that genes transcriptionally modulated by IR were critical to regulate IR-induced biological consequences. Furthermore, eight non-DBS repair genes showed involvement in regulating DSB repair, indicating that successful DSB repair requires both DSB repair mechanisms and non-DSB repair systems. These results reveal that many genes play previously unrecognized roles in multiple DNA repair responses, all of which are required for successful repair of IR-induced damage.
Maintaining vitamin D status without sunlight exposure is difficult without supplementation. This study was designed to better understand interrelationships between periodic vitamin D supplementation ...and immune function in Antarctic workers. The effect of 2 oral dosing regimens of vitamin D supplementation on vitamin D status and markers of immune function was evaluated in people in Antarctica with no UV light exposure for 6 mo. Participants were given a 2000-IU (50 μg) daily (n = 15) or 10,000-IU (250 μg) weekly (n = 14) vitamin D supplement for 6 mo during a winter in Antarctica. Biological samples were collected at baseline and at 3 and 6 mo. Vitamin D intake, markers of vitamin D and bone metabolism, and latent virus reactivation were determined. After 6 mo, the serum 25-hydroxyvitamin D concentration (mean ± SD) increased from 56 ± 17 to 79 ± 16 nmol/L and from 52 ± 10 to 69 ± 9 nmol/L in the 2000-IU/d and 10,000-IU/wk groups, respectively (main effect over time, P < 0.001). Participants with a greater BMI (participant BMI range = 19–43 g/m2) had a smaller increase in 25-hydroxyvitamin D after 6-mo supplementation (P < 0.05). Participants with high serum cortisol and higher serum 25-hydroxyvitamin D were less likely to shed Epstein-Barr virus in saliva (P < 0.05). The doses given raised vitamin D status in participants not exposed to sunlight for 6 mo, and the efficacy was influenced by baseline vitamin D status and BMI. The data also provide evidence that vitamin D, interacting with stress, can reduce risk of latent virus reactivation during the winter in Antarctica.
High amylose starch has many benefits to human health besides diverse industrial applications. Traditional maize grains possess <35% amylose and <5% resistant starch. Here, 48 subtropically adapted ...maize inbreds were evaluated at three locations for various starch attributes. Wide genetic variation for total starch (66.8–74.1%), amylose (1.7–66.2%) and resistant starch (1.4–39.4%) was observed. Minor influence (<5% of total variation) of environment and genotype × environment interactions was observed across the traits. PMI-AML-146 and PMI-AML-147 possessed high amylose (66.2% and 64.2%) and resistant starch (39.4% and 38.0%). At the same time, PMI-WX1 and PMI-WX3 possessed low amylose (1.7% and 3.6%) and resistant starch (1.4% and 1.7%). Amylose was positively correlated with resistant starch (RS, r=0.81) except in a few inbreds. Though CML373 possessed comparatively higher amylose (47.7%), it had a low RS (16.8%). On the contrary, PMI-AML-150 had lower amylose (25.5%) but higher RS (21.4%), suggesting that besides amylose, other factors also play a role in constituting RS. The high amylose and RS inbreds identified here would serve as donors in the breeding programme. This is the first report on genetic variation in amylose and resistant starch in subtropically adapted maize.
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•Amylose (1.7–66.2 %) and resistant starch (1.4–39.4 %) varied greatly among inbreds.•PMI-AML-146 & PMI-AML-147 possessed high amylose (>64 %) & resistant starch (>37 %).•PMI-WX1 and PMI-WX3 possessed low amylose (<4 %) and resistant starch (<2 %).•Amylose showed positive correlation with RS (r = 0.81).
High amylopectin-based waxy maize is popular as food-, feed- and industrial products. Traditional maize is low in amylopectin (∼70–75%), while waxy maize due to the recessive waxy1 (wx1) gene ...possesses ∼95–100% amylopectin. Here, the recessive wx1 gene was introgressed (from MGU-102-wx1) into three elite inbreds that are the parents of two popular sub-tropically adapted maize hybrids viz., HM4 (HKI1105 × HKI323) and HM9 (HKI1105 × HKI1128). BC1F1, BC2F1 and BC2F2 populations were successfully genotyped using gene-based SSR, phi022. The Wx1 gene segregated as per Mendelian inheritance. Background selection using 104–107 SSRs helped in recovering >94% of the recurrent parent genome. MAS-derived inbreds possessed an average of 96.7% amylopectin (range: 95.1–98.7%) compared to 70.5% (range: 69.2–71.6%) among original inbreds. The reconstituted waxy hybrids also recorded significantly higher amylopectin (mean: 98.0%, range: 96.9–98.9%) compared to original hybrids (mean: 69.7%, range: 69.1–70.2%). Waxy hybrids both in yellow and white genetic backgrounds possessed similar grain yield (mean: 5871 kg/ha, range: 5511–6295 kg/ha) with their original versions (mean: 5914 kg/ha, range: 5817–6010 kg/ha) across three locations. These waxy hybrids rich in amylopectin hold significant promise for food security and industrial use. This is the first report of conversion of normal maize hybrids to high amylopectin using genomics-assisted breeding.
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•Traditional maize possesses low amylopectin thus low in food and industrial value.•Recessive wx1 gene was introgressed into two popular white maize hybrids.•Hybrids with wx1 possessed 98.0% amylopectin compared to 69.7% in original hybrids.•Waxy hybrids had similar grain yield with their original versions.•First report of development of waxy maize using genomics-assisted breeding.
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