The noroviruses are important pathogen of epidemic and sporadic gastroenteritis in all age group and show great genetic diversity. The aim of the present study was to describe the prevalence and genetic diversity of noroviruses among children hospitalized with acute sporadic gastroenteritis in Kaohsiung, Taiwan. Fecal samples were collected from hospitalized pediatric patients with sporadic gastroenteritis below age of 18 years during a 2-year period (2007 to 2008). Norovirus RNA was detected by reverse transcription-polymerase chain reaction and comfirmed by sequence analysis. Two different sets of primers were used. Region C primers target shell domain at 5¡¦ end of capsid gene and region D primers target highly variable P2 subdomain at 3¡¦ end of capsid gene. Noroviruses were identified in 5 of 39 (12%) rotavirus-negative specimens using region C primers. Using region D primers only one among these 5 samples could yield PCR product, which showed concordant noroviral genotype. 3 (10%, n=30) specimens from children below age of 5 years tested positive. All these 5 patients had symptoms of vomiting and 3 had fever. All PCR products were sequenced and showed 2 strains of genogroup 1 (G 1.4) and 3 strains of genogroup2 (G 2.4). To our knowledge, this is the first report that demostrated G1.4 genotype norovirus from Taiwan. Norovirus accounted for 10% of sporadic non-bacterial gastroenteritis cases among hospitalized children below 5 years of age in Kaohsiung, Taiwan.

Thesis (M.S.)--University of Tennessee Health Science Center, 2008. / Title from title page screen (viewed on February 24, 2009). Research advisor: Lorraine M. Albritton Ph.D. Document formatted into pages (vii, 81p. : ill.). Vita. Abstract. Includes bibliographical references (p. 65-74).

No description available.

Aging, characterized by a time-dependent functional decline, eventually results in the death of an organism. Unfortunately, this complex biological phenomenon is poorly understood. In order to dissect the molecular changes associated with aging, the identification and molecular characterization of the genes that regulate this universal process is absolutely necessary. The expectation is that the isolated genes potentially have human homologues and can be experimentally analyzed in Drosophila melanogaster in order to determine basic function. In an attempt to find candidate genes that may influence aging, the enhancer trap technique was utilized to identify age-related regulatory elements. The genomic regions surrounding the insertion site of the enhancer trap lines have the potential to be regulated by the characterized enhancer. A previous screen determined the temporal pattern of 180 enhancers trap lines, known as DJ lines. Many of these lines demonstrated an expression pattern that was associated with age. Several of the genes within the nearby genomic regions of six sequenced DJ lines, DJ695, DJ710, DJ849, DJ767, DJ761 and DJ694, were chosen for transcript quantification. Prior to gene quantification, reverse transcription, an essential step in the experimental procedure, was assessed for the error it incorporated into quantification. Specifically, an exogenous molecule was used to ensure that unsuccessful reverse transcription reactions had the potential to be identified and, soon after, discarded. This was achieved through the use of a spike RNA molecule, Luciferase. Luciferase was shown to be a diagnostic tool that can be used in determining reverse transcription efficiency. Eight genes were chosen from the aforementioned DJ lines and quantitative PCR revealed that the natural regulation of some genes were comparable to the, previously obtained, expression pattern of the enhancer trap line. Although the expression of other genes did not correlate to that of the enhancer trap lines, all genes exhibited expression patterns that were age-associated. The known functions of these candidate genes and the relevant homologues are discussed. These findings validate the use of the enhancer trap technique in the identification of candidate genes involved in the aging process. / Thesis (Master, Biology) -- Queen's University, 2009-02-09 10:59:16.871

Thesis (M.S.)--Florida State University, 2005. / Advisor: Dr. Ross Ellington, Florida State University, College of Arts and Sciences, Dept. of Biological Science. Title and description from dissertation home page (viewed June 8, 2005). Document formatted into pages; contains xiii, 55 pages. Includes bibliographical references.

Thesis (Ph. D.)--University of Alabama at Birmingham, 2007. / Title from first page of PDF file (viewed June 23, 2008). Includes bibliographical references.

Le VIH-1 est un rétrovirus contenant deux copies de son génome ARN simple brin positif. Dans la cellule, son ARN est rétrotranscrit en ADN puis cet ADN est intégré dans le génome cellulaire. L'ADN viral est ensuite transcrit en ARN dont la moitié évite l'épissage. Dans le cytoplasme, cet ARN possède une double fonctionnalité : il sert d'ARNm pour la synthèse des protéines Gag et Gag-Pol et d'ARN génomique (ARNg) qui est encapsidé sous forme de dimère. Malgré sa faible abondance dans la cellule, l'ARNg est préférentiellement encapsidé. Cependant, il a été montré par nous et d'autres que d'autres ARN non-génomiques peuvent également être spécifiquement encapsidés, tels que des ARN viraux épissés et certains ARN cellulaires, bousculant ainsi le dogme d'une spécificité unique pour l'ARNg. Nous avons montré que le VIH-1 contrôlerait l'incorporation des ARN viraux, épissés et non épissés, par un mécanisme de compétition, impliquant des facteurs communs. En revanche, les ARN cellulaires 7SL et U6 semblent encapsidés par des mécanismes indépendants. Afin d'identifier les déterminants qui régissent la spécificité d'encapsidation, nous avons testé le rôle de la nucléocapside (NC) qui est fortement impliquée dans l'encapsidation. Nous montrons ici que de façon tou t à fait inattendue, des mutations de NC conduisent à l'encapsidation d'ADN et augmentent également l'encapsidation d'ARN viraux épissés. Nous avons ensuite cartographié les déterminants de la région 5' UTR de l'ARNg et déterminé que la tige-boucle polyA et la région PBS sont impliquées dans l'encapsidation des ARN viraux épissés. Ce travail apporte une meilleure compréhension de la spécificité d'encapsidation, primordiale pour la mise au point de thérapies géniques utilisant des vecteurs lentiviraux. / HIV-1 is a retrovirus containing two copies of its single stranded positive RNA genome. In the cell, its RNA is reverse transcribed into DNA and this DNA is integrated into the cellular genome. The viral DNA is then transcribed into RNA. One moiety of this RNA pool escapes splicing. Once in the cytoplasm, this RNA has a double function: it serves as mRNA for synthesis of Gag and Gag-Pol proteins and as genomic RNA (gRNA) that is packaged as a dimer. Despite its low abundance in the cell, the gRNA is preferentially encapsidated into virions. However, it has been shown by us and others that other non-genomic RNA can be specifically packaged, such as spliced viral RNA and some cellular RNA, thus shaking up the dogma of a unique specificity for the gRNA. We have shown that HIV-1 might control the incorporation of the spliced and unspliced RNA by a mechanism of competition, involving common factors. In contrast, cellular RNA 7SL and U6 seem encapsid ated through independent mechanisms. To identify the determinants that govern the specificity of encapsidation, we tested the role of the nucleocapsid (NC) which is strongly involved in the packaging. Here we show that unexpectedly, mutations of NC lead to the encapsidation of DNA and also increase the encapsidation of spliced viral RNA. We then mapped the determinants in the 5' UTR of the gRNA and determined that the polyA stem-loop and the PBS region are involved in the encapsidation of the spliced viral RNA. This work provides a better understanding of the specificity of encapsidation that is crucial for the development of gene therapy using lentiviral vectors.

Crimean-Congo Hemorrhagic Fever (CCHF) is a worldwide tick-borne disease that originally belongs to the Bunyaviridae family, the genus Nairovirus. In addition to infection from ticks, humans become infected if any contact with infected blood or tissue material occurs. To study the disease, several methods such as real-time Polymerase Chain Reaction, enzyme-linked immunosorbent assay and Immunofluorescence assay are used for detection of the virus. All viruses in Bunyaviridae consists of three single stranded RNA sequences, the small, the medium and the large segment, that encode for the nucleocapsid protein, the glycoproteins, GN and GC, and the RNA-dependent RNA polymerase, respectively. The main purpose of this study was to express the M RNA segment´s glycoproteins, GN and GC. By using the reverse transcription reaction, the cDNA was synthesized from vRNA and the M RNA sequence was amplified using Phusion DNA-polymerase. In the storage vector, pcDNA3.1/V5-His-TOPO, the insert was ligatured followed by transformation into Escherichia coli. Restriction digestion was made with specific enzymes that cut out the insert. In the second ligation and transformation two different expression vectors (pTM1/pI.18) was used. After observation of the gel analysis from the test-PCR, an insert in the expression vector was shown.

Celiac disease (CD) is a common heritable immune related disorder where chronic inflammationof the small intestine is induced by the ingestion of gluten. The immune response leads to theinflammation and flattening of intestinal mucosa due to the damaged villi and thus results indefects in the absorption of nutrients. This defect can affect any organ or body system and exposeto the risk of lifelong complications such as cancer, autoimmune diseases and other complexdiseases. Now a day, celiac disease is becoming one of the well-studied models of complexdisorders.The PI3K- FOX signaling pathway is activated by many regulators and growth factors and playsa key role in cell cycle. Two components of this pathway, INSR and FOX, play crucial roles indiverse aspects of embryogenesis from the initial tissue genesis up to organ formation. INSR andFOX take part in development, differentiation, proliferation, apoptosis and stress resistance aswell as metabolism. SNP´s could affect the expression of neighboring genes. These SNP´s areshown to be as eQTLs, genomic loci that regulate the expression of genes. The aim of this studywas to detect and quantitate the expression of INSR and certain FOX genes in celiac disease.Quantitative real time PCR (QPCR) was used to analyze the expression of INSR, FOXO1,FOXO4 and FOXD3 genes in 38 celiac cases and 50 control samples. Three reference genesACTB, EPCAM and PGK1 were tested for their expression stability and their average was used inthe normalization procedure. Gene expression results were analyzed using the ΔCt method. Theexpression of INSR, FOXO1, FOXO4 and FOXD3 were described as their fold change in CDcompared to normal non-celiac mucosa. Our results indicated that FOXO4 and INSR wereexpressed less by 0.60 fold and FOXO1 was expressed less by 0.23 fold in CD samples. Theresults are preliminary and further studies will be needed to confirm if these findings are a resultof the intestinal inflammation in CD or if these genes are partly driving the disease itself.

A simple anaerobic digestion model has been developed for a continuously-stirred tank reactor (CSTR), which links the specific biogas production rate to the food/microorganism ratio (F/M). The model treats the various microbial populations involved in the sequential biological processes involved in anaerobic digestion as a composite and links the entire biomass specific growth rate directly to the specific biogas production rate. The model was calibrated by determining the specific gas production rate for a range of F/M values using a municipal wastewater seed sludge. The model predictions for steady-state biogas production rates were compared to observed biogas production and volatile solids destruction results from three laboratory-scale anaerobic digesters that were operated at hydraulic retention times of 10, 15, and 20 days. The F/M model results were shown to agree with reactor biogas output for 10, 15, and 20 day hydraulic retention times to within 5.0%, 14.3%, and 9.5%, respectively. A commercial wastewater treatment plant model, BioWin 3, was also used to model anaerobic digestion as a comparison. Agreement for the BioWin 3 model results, as compared to the 10, 15, and 20-day hydraulic retention time reactors, was within 66.2%, 114.1%, and 105.1%, respectively. In all cases the BioWin 3 model over-predicted biogas output as compared to the reactors. A molecular biology method called RT-RiboSyn was developed to measure the specific growth rate of microbial populations. RT-RiboSyn, is an ex situ method that utilizes a reverse transcription and primer extension (RT&PE) method to analyze the rRNA extracted from a time series of samples treated with chloramphenicol. The method measures the rate of ribosome synthesis over time through the increase in precursor 16S rRNA (pre16S rRNA) relative to the mature 16S rRNA (16S rRNA). A single fluorescently labeled primer that targets an interior region of both pre16S and 16S rRNA for a distinct population is used to generate two pools of reverse transcription product. The ratio of pre16S and 16S rRNA is then determined by separating these pools by length using capillary electrophoresis, and measuring the fluorescent intensity of each pool of fragments. Results from three different log growth cultures of Acinetobacter calcoaceticus indicate that RT-RiboSyn, as compared to spectrophotometer readings, was able to predict specific growth rates within -3.1% to 10% and -3.3% to 21.0% when using a primer targeting Eubacteria and Acinetobacter, respectively. The RT-RiboSyn results from a stationary phase culture predicted no growth and possible 16S rRNA degradation. Further work was completed to determine whether the RiboSyn method would successfully measure growth rates of specific microbial populations in environmental samples. The first of these was activated sludge from a high-purity oxygen system in a wastewater treatment facility located in Tampa, Florida. The organism targeted was the Acinetobacter genus, which was shown to be prevalent via fluorescence in situ hybridization results. RT-RiboSyn results indicated that growth was not measureable for the Acinetobacter present in the system; however, since the sludge was taken at the end of the process, Acinetobacter may have been in stationary phase when the samples were collected. Attempts were made to apply the method to methanogens in both pure culture and anaerobic digester sludge samples. An analysis of samples of RNA from Methanosarcina barkeri indicated that the presence of 16S rRNA could be measured; however, capillary electrophoresis instrument limitations prevented the detection of pre16S rRNA fragments. Additional testing of anaerobic digester sludge for both bacterial and Archaeal population was successful for detecting 16S rRNA and possibly precursor 16S rRNA fragments of a variety of lengths. However, specific growth rates could not be determined for the Archaea present in these samples, either due to capillary electrophoresis limitations or very slow growth rates. The results show that the RT-RiboSyn method is applicable to pure cultures; however, a modification of the method is needed to overcome the limitations apparent in populations with low specific growth rates.