Biomedical subjects

Y Tang

Publications and source records attributed to Y Tang.

At least 19 recordsLinked to original sources

PubMed2007

Regulation of oxidant-induced intestinal permeability by metalloprotease-dependent epidermal growth factor receptor signaling.

Inflammatory bowel disease (IBD) affects more than 1 million Americans with more than 30,000 new cases diagnosed each year. IBD increases patient morbidity and susceptibility to colorectal cancer, yet its etiology remains unknown. Current models identify two key determinants of IBD pathogenesis: hyperpermeability of the gut epithelial barrier to bacterial products and an abnormal immune response to these products. Two factors seem critical for hyperpermeability: oxidant-induced stress and proinflammatory cytokines (e.g., tumor necrosis factor-alpha). The aim of this study was to investigate the role of oxidant stress-mediated transactivation of the epidermal growth factor receptor (EGFR) in intestinal hyperpermeability. This study used the Caco-2 human colonic epithelial cell in vitro model of intestinal epithelium. Cells were grown on inserts for permeability and signaling studies and glass coverslips for microscopy studies. show that oxidant-induced intestinal hyperpermeability can be blocked by specific inhibitors of the EGFR, tumor necrosis factor convertase (TACE) metalloprotease, transforming growth factor (TGF)-alpha, and mitogen-activated protein kinases, especially extracellular signal-regulated kinase 1/2. We also show that oxidant initiates these signaling events, in part by causing translocation of TACE to cell-cell contact zones. In this study, our data identify a novel mechanism for oxidant-induced intestinal hyperpermeability relevant to IBD. We propose a new intestinal permeability model in which oxidant transactivates EGFR signaling by activation of TACE and cleavage of precursor TGF-alpha. These data could have a significant effect on our view of IBD pathogenesis and provide new therapeutic targets for IBD treatment.

ADAM Proteins↗

PubMed2006

Directing adenovirus across the blood-brain barrier via melanotransferrin (P97) transcytosis pathway in an in vitro model.

Adenovirus serotype 5 (Ad5) is widely used in the development of gene therapy protocols. However, current gene therapy strategies involving brain are mostly based on intra-cranial injection. A major obstacle for systemically administered vectors to infect brain tissue is the blood-brain barrier (BBB). One strategy to cross the BBB is transcytosis, a transcellular transport process that shuttles a molecule from one side of the cell to the other side. Recently, melanotransferrin (MTf)/P97 was found to be able to cross the BBB and accumulate in brain. We thus hypothesize that re-directing Ad5 vectors to the MTf transcytosis pathway may facilitate Ad5 vectors to cross the BBB. To test this hypothesis, we constructed a bi-specific adaptor protein containing the extracellular domain of the coxsackie-adenovirus receptor (CAR) and the full-length melanotransferrin (sCAR-MTf), and investigated its ability to re-direct Ad5 vectors to the MTf transcytosis pathway. We found this adaptor protein could re-direct Ad5 to the MTf transcytosis pathway in an in vitro BBB model, and the transcytosed Ad5 viral particles retained their native infectivity. The sCAR-MTf-mediated Ad5 transcytosis was temperature- and dose dependent. In addition, we examined the directionality of sCAR-MTf-mediated Ad5 transcytosis, and found the efficiency of apical-to-basal transcytosis was much higher than that of basal-to-apical direction, supporting a role of this strategy in transporting Ad5 vectors towards the brain. Taken together, our study demonstrated that re-directing Ad5 to the MTf transcytosis pathway could facilitate gene delivery across the BBB.

Adenoviridae↗

PubMed2006

Expression of the novel co-stimulatory molecule B7-H4 by renal tubular epithelial cells.

Crosstalk between T cells and renal tubular epithelial cells (TECs) in the pathogenesis of tubular lesions, the most important sign of progressive renal diseases, has not been clarified. Previous work has shown that TECs harbor co-stimulatory signals that promote T-cell activation, which induces tubular lesions. Nevertheless, the expression and functional role of B7-H4, a recently identified co-stimulatory ligand of the B7 superfamily, in pathologic human kidneys is unclear. We investigated the expression of B7-H4 on cryostat renal biopsies from patients with idiopathic membranous nephropathy (n=20), immunoglobulin A nephropathy (n=19), lupus nephritis (n=16), and acute renal allograft rejection (n=15) using immunohistochemistry. In addition, we also analyzed TEC-associated B7-H4 in the regulation of T-cell activation. Immunohistological staining revealed that B7-H4 antigen is restricted to tubular epithelium and that the protein is prominent in sections with severe tubular lesions, although no correlation was observed between tubular B7-H4 expression and levels of serum creatinine, serum urea nitrogen concentration, and 24-h proteinuria in each type of nephropathy. In vitro, mixed lymphocyte reactions revealed that TEC-related B7-H4 promotes cytokine (interleukin-2 and interferon-gamma) production and proliferation of co-cultured T cells. Interestingly, the secretion of interleukin-2 by C10 T cell hybridomas also increased when C10 cells were co-cultured with the B7-H4-transgenic murine TEC line, 3M-1-secreting tubular epithelial cells (MCT) in the presence of the antigen hen egg lysozyme. Our results clearly show that TEC-associated B7-H4 induces T-cell activation and we propose that B7-H4 is a potential activator that promotes tubular lesion.

Animals↗

PubMed2006

The relationship between cerebral Alzheimer's disease pathology and odour identification in old age.

BACKGROUND: Olfactory dysfunction is common in old age, but its basis is uncertain. OBJECTIVE: To test the hypothesis that difficulty in identifying odours in old age is related to the accumulation of Alzheimer's disease pathology. METHODS: As part of the Rush Memory and Aging Project, participants completed the 12-item Brief Smell Identification Test, a standard measure of odour identification. During a mean (standard deviation (SD)) of 2.2 (1.2) years of follow-up (range 0.2-4.9), 166 people died, with brain autopsies performed on 129 (77.7%) people and neuropathological examinations completed on 77 (mean (SD) age at death 87.5 (5.9) years; median postmortem interval 6.1 h). From a uniform postmortem examination of multiple brain regions, summary measures of plaque and tangle pathology were derived on the basis of silver staining, and those of amyloid beta burden, tangle density and Lewy bodies on the basis of immunohistochemistry. RESULTS: Odour identification performance ranged from 0 to 12 correct (mean (SD) 8.0 (2.6)). In analyses adjusted for age, sex and education, a composite measure of plaques and tangles accounted for >12% of the variation in odour identification. The association remained after controlling for dementia or semantic memory. Density of tau tangles was inversely related to odour identification. A similar effect for amyloid burden was attenuated after controlling for tangles. The association with odour identification was robust for tangles in the entorhinal cortex and CA1/subiculum area of the hippocampus, but not for tangles in other cortical sites. Lewy bodies, identified in 12.5%, were not related to odour identification, probably partly due to to their relative infrequency. CONCLUSION: The results suggest that difficulty in identifying familiar odours in old age is partly due to the accumulation of neurofibrillar pathology in central olfactory regions.

Aged↗

PubMed2006

Mild cognitive impairment: risk of Alzheimer disease and rate of cognitive decline.

OBJECTIVE: To examine the extent to which persons with mild cognitive impairment (MCI) have an increased risk of Alzheimer disease (AD) and a more rapid rate of decline in cognitive function compared to similar persons without cognitive impairment. METHOD: Participants were 786 community-based persons (221 with MCI and 565 without cognitive impairment) from the Rush Memory and Aging Project, an ongoing longitudinal clinical-pathologic study of common chronic conditions of old age. All participants underwent detailed annual clinical and neuropsychological evaluations. The authors examined the risk of incident AD and rate of change in global cognitive function among persons with MCI and those without cognitive impairment; all statistical models controlled for age, sex, and education. RESULTS: Over an average of 2.5 years of follow-up, 57 persons with MCI (25.8%) developed AD, a rate 6.7 times higher than those without cognitive impairment. In addition, persons with MCI declined considerably more rapidly each year on a measure of global cognitive function than those without cognitive impairment. CONCLUSIONS: Mild cognitive impairment is associated with a greatly increased risk of incident Alzheimer disease and a more rapid rate of decline in cognitive function.

Age Factors↗

PubMed2006

Doppel-induced apoptosis and counteraction by cellular prion protein in neuroblastoma and astrocytes.

Expression of a prion-like protein, doppel, induces apoptosis-like changes in cerebellar neuronal granule and Purkinje cells of prion-knockout mice and this effect can be rescued by re-introduction of cellular prion. Since most of those studies were done in transgenic mice, in the present study, we have established a murine neuro-2a cell line and the primary rat adult reactive astrocyte model for studying doppel-induced apoptosis and possible prion counteraction. We demonstrate that expression of doppel in neuro-2a cells causes apoptosis, during which DNA fragmentation occurs as visualized by terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling staining and other intracellular changes characteristic of apoptosis are observed in the electron microscope. Using immunoblot analyses, we further demonstrate that doppel expression activates caspase-10 as well as caspase-3, but does not activate caspase-9. Addition of purified doppel to cultures of neuro-2a cells and the primary astrocytes causes similar apoptotic changes. Significantly, apoptosis induced by doppel is enhanced when cellular prion protein is depleted by RNA interference, suggesting a protective effect of cellular prion against doppel-induced apoptosis. The antagonistic interaction between cellular prion and doppel appears to involve direct protein-protein interaction possibly on cell membrane as cellular prion and doppel physically interact with each other and co-localize on cell membranes. Together, our data show that doppel induces apoptosis in neuroblastoma neuro-2a and rat primary astrocytes via a caspase-10 mediated pathway and that this effect is counteracted by cellular prion through direct interaction with doppel possibly on cell membrane.

Animals↗

PubMed2006

A genome-scale assessment of peripheral blood B-cell molecular homeostasis in patients with rheumatoid arthritis.

OBJECTIVE: While rheumatoid arthritis (RA) is considered a prototypical autoimmune disease, the specific roles of B-cells in RA pathogenesis is not fully delineated. METHODS: We performed microarray expression profiling of peripheral blood B-cells from RA patients and controls. Data were analysed using differential gene expression analysis and 'gene networking' analysis (characterizing clusters of functionally inter-relelated genes) to identify both regulatory genes and the pathways in which they participate. Results were confirmed by quantitative real-time polymerase chain reaction and by measuring the levels of 10 serum cytokines involved in the pathways identified. RESULTS: Genes regulating and effecting the cell-cycle, proliferation, apoptosis, autoimmunity, cytokine networks, angiogenesis and neuro-immune regulation were differentially expressed in RA B-cells. Moreover, the serum levels of several soluble factors that modulate these pathways, including IL-1beta, IL-5, IL-6, IL-10, IL-12p40, IL-17 and VEGF were significantly increased in this cohort of RA patients. CONCLUSIONS: These results outline aspects of the multifaceted role B-cells play in RA pathogenesis in which immune dysregulation in RA modulates B-cell biology and thereby contributes to the induction and perpetuation of a pathogenic humoral immune response.

Adult↗

PubMed2006

Critical interactions between TGF-beta signaling/ELF, and E-cadherin/beta-catenin mediated tumor suppression.

Inactivation of the transforming growth factor-beta (TGF-beta) pathway occurs often in malignancies of the gastrointestinal (GI) system. However, only a fraction of sporadic GI tumors exhibit inactivating mutations in early stages of cancer formation, suggesting that other mechanisms play a critical role in the inactivation of this pathway. Here, we show a wide range of GI tumors, including those of the stomach, liver and colon in elf+/- and elf+/- / Smad4+/- mutant mice. We found that embryonic liver fodrin (ELF), a beta-Spectrin originally identified in endodermal stem/progenitor cells committed to foregut lineage, possesses potent antioncogenic activity and is frequently inactivated in GI cancers. Specifically, E-cadherin accumulation at cell-cell contacts and E-cadherin-beta-catenin-dependent epithelial cell-cell adhesion is disrupted in elf+/- / Smad4+/- mutant gastric epithelial cells, and could be rescued by ectopic expression of full-length elf, but not Smad3 or Smad4. Subcellular fractionation revealed that E-cadherin is expressed mainly at the cell membrane after TGF-beta stimulation. In contrast, elf+/- / Smad4+/- mutant tissues showed abnormal distribution of E-cadherin that could be rescued by overexpression of ELF but not Smad3 or Smad4. Our results identify a group of common lethal malignancies in which inactivation of TGF-beta signaling, which is essential for tumor suppression, is disrupted by inactivation of the ELF adaptor protein.

Animals↗

PubMed2006

Action mechanisms and structure-activity relationships of PI3Kgamma inhibitors on the enzyme: a molecular modeling study.

Action mechanisms of four types of PI3Kgamma inhibitors were investigated on the ligand-binding pocket (LBP) of PI3Kgamma with molecular modeling method. At first five compounds whose complex structures with PI3Kgamma were available experimentally were used to validate the reliability of docking program Autodock3.0. The results demonstrated that the program could reproduce the bound conformations of those compounds in crystal structures. Then the program was used to dock all the four types of PI3Kgamma inhibitors into the LBP of the enzyme. The predicted activities of these compounds were in agreement with their experimental activities, and a pharmacophore model was hence derived for these compounds, which consisted of one hydrophobic portion flanked by two symmetric hydrophilic portions. Furthermore, the structure-activity relationships of PI3Kgamma inhibitors were elucidated and the activity differences between them were discussed.

Adenosine Triphosphate↗

PubMed2006

RING finger-dependent ubiquitination by PRAJA is dependent on TGF-beta and potentially defines the functional status of the tumor suppressor ELF.

In gastrointestinal cells, biological signals for transforming growth factor-beta (TGF-beta) are transduced through transmembrane serine/threonine kinase receptors that signal to Smad proteins. Smad4, a tumor suppressor, is often mutated in human gastrointestinal cancers. The mechanism of Smad4 inactivation, however, remains uncertain and could be through E3-mediated ubiquitination of Smad4/adaptor protein complexes. Disruption of ELF (embryonic liver fodrin), a Smad4 adaptor protein, modulates TGF-beta signaling. We have found that PRAJA, a RING-H2 protein, interacts with ELF in a TGF-beta-dependent manner, with a fivefold increase of PRAJA expression and a subsequent decrease in ELF and Smad4 expression, in gastrointestinal cancer cell lines (P < 0.05). Strikingly, PRAJA manifests substantial E3-dependent ubiquitination of ELF and Smad3, but not Smad4. Delta-PRAJA, which has a deleted RING finger domain at the C terminus, abolishes ubiquitination of ELF. A stable cell line that overexpresses PRAJA exhibits low levels of ELF in comparison to a Delta-PRAJA stable cell line, where ELF expression is high compared to normal controls. The alteration of ELF and/or Smad4 expression and/or function in the TGF-beta signaling pathway may be induced by enhancement of ELF degradation, which is mediated by a high-level expression of PRAJA in gastrointestinal cancers. In hepatocytes, half-life (t(1/2)) and rate constant for degradation (k(D)) of ELF is 1.91 h and 21.72 min(-1) when coupled with ectopic expression of PRAJA in cells stimulated by TGF-beta, compared to PRAJA-transfected unstimulated cells (t(1/2) = 4.33 h and k(D) = 9.6 min(-1)). These studies reveal a mechanism for tumorigenesis whereby defects in adaptor proteins for Smads, such as ELF, can undergo degradation by PRAJA, through the ubiquitin-mediated pathway.

Animals↗

PubMed2006

Integration of abscisic acid signalling into plant responses.

The phytohormone abscisic acid (ABA) plays a major role as an endogenous messenger in the regulation of plant's water status. ABA is generated as a signal during a plant's life cycle to control seed germination and further developmental processes and in response to abiotic stress imposed by salt, cold, drought, and wounding. The action of ABA can target specifically guard cells for induction of stomatal closure but may also signal systemically for adjustment towards severe water shortage. At the molecular level, the responses are primarily mediated by regulation of ion channels and by changes in gene expression. In the last years, the molecular complexity of ABA signal transduction surfaced more and more. Many proteins and a plethora of "secondary" messengers that regulate or modulate ABA-responses have been identified by analysis of mutants including gene knock-out plants and by applying RNA interference technology together with protein interaction analysis. The complexity possibly reflects intensive cross-talk with other signal pathways and the role of ABA to be part of and to integrate several responses. Despite the missing unifying concept, it is becoming clear that ABA action enforces a sophisticated regulation at all levels.

Abscisic Acid↗

PubMed2006

Changes and relations of circulating visfatin, apelin, and resistin levels in normal, impaired glucose tolerance, and type 2 diabetic subjects.

Visfatin and apelin are two novel adipocyte- secreted hormone proposed to link obesity with insulin resistance. In this study we investigated whether plasma visfatin and apelin levels were altered in normal, impaired glucose tolerance, and type 2 diabetic subjects. We also assessed the association between plasma visfatin, or apelin and body composition, metabolic parameters, and resistin concentrations in these subjects. The visfatin levels of fasting and 2-h post-glucose load were found to be significantly decreased in diabetics compared with the controls ( P<0.05). In contrast, basal apelin levels were significantly increased in the IGT and diabetic subjects compared with the controls ( P<0.05 and P<0.01). The apelin levels of 2-h post-glucose load were significantly higher than the basal levels in every group (all P<0.05). Fasting plasma visfatin was found to correlate positively and significantly with BMI, WHR, and fasting plasma resistin, but negatively with HbA1c and 2 h OGTT glucose. Multiple regression analysis showed that WHR, HbA1c, 2 h OGTT glucose were independent related factors influencing plasma visfatin levels. Fasting plasma apelin levels correlated positively with HOMA-IR, BMI, TC, LDL-C, FBG and Fasting plasma insulin. Multiple regression analysis also showed that HOMA-IR, BMI, and TC were independent related factors influencing plasma apelin levels. The present work indicates the potential link of visfatin and apelin with the pathogenesis of insulin resistance and T2DM.

Adult↗

PubMed2006

Novel IRF6 mutations in Chinese patients with Van der Woude syndrome.

Van der Woude syndrome (VWS) (OMIM 119300) is a dominantly inherited, developmental disorder that is characterized by pits and/or sinuses of the lower lip and a cleft lip and/or cleft palate. Mutations in the interferon regulatory factor 6 gene (IRF6) have been recently identified in patients with VWS, with more than 60 mutations reported. However, the VWS phenotype, IRF6 mutation genotypes, and their interrelationships in Chinese VWS patients have not been studied. Here, we report 11 Chinese families with variable clinical phenotypes of VWS and identified mutations in all patients. Of the 11 mutations, 8 appeared to be novel: CC5.6GT, T342A, 566delA, C748T, C756A, C989A, C1209G, and 1316delT. Seven mutations caused a change or loss of the IRF6 domain. The marked phenotypic variation may be caused by the action of certain modifier genes on IRF6 function.

Adult↗

PubMed2006

[Comparative study on clinical features of 215 patients with acute and subacute severe hepatitis.].

BACKGROUND: To analyze the clinical features of acute severe hepatitis (ASH) and subacute severe hepatitis (SSH) by comparison analysis. METHODS: The clinical features of 215 cases with ASH and SSH from 1995 to 2005 were retrospectively analyzed by using chi-test, t-test and regression analysis. RESULTS: The most common cause of ASH or SSH was hepatitis B virus (HBV) infection (>30%). Anti-tuberculosis therapy was the main cause of drug-induced ASH or SSH. The incidences of hepatic encephalopathy were 78.13% and 43.05% respectively (P less than 0.001). Prothrombin activity was less than 20% and less than 30%, respectively in cases with ASH and SSH. The most common complications of ASH were hepatic encephalopathy, imbalance of electrolyte and cerebral edema, and that of SSH were ascites, imbalance of electrolyte and hepatic encephalopathy. Prothrombin time, ratio of WBC and neutrophil count were positively related to case fatality of ASH or SSH. Prothrombin activity and total cholesterol (TC) were negatively correlated with fatality of ASH or SSH. Total bilirubin, blood ammonia and creatinine were also positively correlated with fatality of SSH; chlolinesterase, triglyceride, PLT, albumin were negatively correlated with fatality of SSH. CONCLUSION: ASH and SSH are two different entities because of obvious differences in the ages of suffering, the incidence and occurring time of hepatic encephalopathy, abnormality of function of blood coagulation and the marker for predicting prognosis. Severe abnormality of function of blood coagulation is an important marker with high sensitivity and specificity in ASH or SSH patients without hepatic encephalopathy.

Hepatitis↗

PubMed2005

Hallucinations, cognitive decline, and death in Alzheimer's disease.

The relation of psychotic symptoms to cognitive decline and mortality in Alzheimer's disease (AD) was examined during a mean of 2.2 years in 478 persons selected from clinical settings. Psychotic symptoms were ascertained at baseline and cognition was assessed semiannually with nine tests from which a global measure was formed. In analyses that controlled for age, sex, race, and education, hallucinations (29.6%), especially visual ones, were associated with more rapid global cognitive decline and increased mortality, even after controlling for baseline level of cognition and use of antipsychotic medication, and the association with mortality increased with higher level of education. Delusions and misperceptions were not strongly related to cognitive decline or mortality. The results suggest that hallucinations in Alzheimer's disease, particularly visual ones, are associated with more rapid progression.

Aged↗

PubMed2005

Effects of alveolar dead-space, shunt and V/Q distribution on respiratory dead-space measurements.

BACKGROUND: Respiratory dead-space is often increased in lung disease. This study evaluates the effects of increased alveolar dead-space (Vd(alv)), pulmonary shunt, and abnormal ventilation perfusion ratio (/) distributions on dead-space and alveolar partial pressure of carbon dioxide (Pa(co(2))) calculated by various methods, assesses a recently published non-invasive method (Koulouris method) for the measurement of Bohr dead-space, and evaluates an equation for calculating physiological dead-space (Vd(phys)) in the presence of pulmonary shunt. METHODS: Pulmonary shunt, / distribution and Vd(alv) were varied in a tidally breathing cardiorespiratory model. Respiratory data generated by the model were analysed to calculate dead-spaces by the Fowler, Bohr, Bohr-Enghoff and Koulouris methods. Pa(co(2)) was calculated by the method of Koulouris. RESULTS: When Vd(alv) is increased, Vd(phys) can be recovered by the Bohr and Bohr-Enghoff equations, but not by the Koulouris method. Shunt increases the calculated Bohr-Enghoff dead-space, but does not affect Fowler, Bohr or Koulouris dead-spaces, or Vd(phys) estimated by the shunt-corrected equation if pulmonary artery catheterization is available. Bohr-Enghoff but not Koulouris or Fowler dead-space increases with increasing severity of / maldistribution. When alveolar Pco(2) is increased by any mechanism, Pa(co(2)) calculated by Koulouris' method does not agree well with average alveolar Pco(2). CONCLUSIONS: Our studies show that increased pulmonary shunt causes an apparent increase in Vd(phys), and that abnormal / distributions affect the calculated Vd(phys) and Vd(alv), but not Fowler dead-space. Dead-space and Pa(co(2)) calculated by the Koulouris method do not represent true Bohr dead-space and Pa(co(2)) respectively, but the shunt-corrected equation performs well.

Adult↗

PubMed2005

Sensitive detection of human papillomavirus in cervical, head/neck, and schistosomiasis-associated bladder malignancies.

We assayed for the presence of human papilloma virus (HPV) DNA in serum and/or peripheral blood fraction (PBF) of individuals with cervical, head/neck, or bladder cancer due to schistosomiasis. Using mass spectroscopy coupled with competitive PCR, HPV DNA was detected at the individual molecule level by using "MassARRAY" assays. The resultant sensitivity was superior to real-time fluorescent PCR-based assays, while specificity was maintained. Our principal findings were: (i) Virtually all tested cervical cancers and schistosomiasis-associated bladder cancers, and a plurality of head/neck cancers, are associated with HPV DNA in the tumor. (ii) All 27 bladder cancers due to schistosomiasis were associated with the presence of HPV-16 DNA, which can be detected in tumor and serum but not in PBF. In contrast, no serum HPV-16 DNA signal was detected in seven individuals with schistosomiasis-associated bladder cancers after surgical removal of the tumor. (iii) Among the head/neck cancers we studied, anterior tumors were more often associated with HPV DNA in tumor, serum, and/or PBF than posterior tumors. (iv) In cervical cancer, where all tumors contain HPV DNA, viral DNA could be detected often in serum and/or PBF. Further, HPV-16 DNA was detected in serum and/or PBF of most patients with untreated high-grade cervical dysplasia but disappeared if the dysplasia was eliminated. The sensitive, specific, and quantitative MassARRAY technique should make it feasible to monitor cancer occurrence and treatment and recurrence of malignancies and dysplasias associated with HPV DNA.

DNA Primers↗

PubMed2005

Development of a ssRNA internal control template reagent for a multiplex RT-PCR to detect turkey astroviruses.

Turkey astrovirus (TAstV), a positive sense single-stranded RNA (ssRNA) virus, is an important causative agent of enteritis of poults. The detection and diagnosis of astroviruses have been mainly dependent on electron microscopy (EM). However, EM is not very sensitive. Reverse transcription-polymerase chain reaction (RT-PCR) has high specificity and sensitivity. Thus, monoplex RT-PCR and multiplex RT-PCR for detection of TAstVs were developed in our laboratory. RT-PCR could be adversely affected by many factors, which would result in lowering the sensitivity of the reaction. To minimize this pitfall of RT-PCR, a ssRNA internal control (IC) template reagent containing two sets of primer sequences (SRV and AFCP) specific to the capsid region of TAstV genomes was developed, and applied to the multiplex RT-PCR. Sixty-four fecal specimens from 2-week-old poults with enteritis were tested using the multiplex RT-PCR with the IC serving as a co-amplification template. An overall test inhibition rate of 12.5% was found for the RT-PCR, which can be used to decrease the false-negative rate. The approach to the generation of the IC developed in this study is simple, convenient and productive, and can be used as a universal protocol to generate a ssRNA IC template reagent for RT-PCR.

Animals↗