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Polymerase Chain Reaction Techniques (polymerase + chain_reaction_techniques)
Selected AbstractsPrevalence and potential link between E. coli O157:H7 isolated from drinking water, meat and vegetables and stools of diarrhoeic confirmed and non-confirmed HIV/AIDS patients in the Amathole District , South AfricaJOURNAL OF APPLIED MICROBIOLOGY, Issue 2 2008B.O. Abong'o Abstract Aim:, The current study investigated the prevalence and molecular relatedness between Escherichia coli O157:H7 isolated from water, meat and meat products and vegetables and from stools of confirmed and non-confirmed Human Immune Virus/Acquired Immunodeficiency Syndrome (HIV/AIDS) patients with diarrhoea. Methods and Results:, Culture-based and polymerase chain reaction techniques were used to identify E. coli O157:H7. Thirty-five per cent of meat products, 25·5% of water, 21·7% of vegetables as well as 56·5% and 43·5% of stools of confirmed and non-confirmed HIV/AIDS patients, respectively, were presumptively positive with E. coli O157. Molecular results indicated that 10·3%, 8·6% and 7·8% of the vegetables, water and meat products examined carried E. coli O157:H7, which had homologous fliCH7, rfbEO157 and eaeA genetic loci to the genes of some E. coli O157:H7 isolated from 12·2% and 8·8% of the stools of confirmed and non-confirmed HIV/AIDS patients, respectively. Conclusions:, Water, meat and meat products and vegetables are potential sources of E. coli O157:H7 that are potentially capable of causing diarrhoea in humans especially HIV/AIDS patients. Significance and Impact of the Study:, Great care should be exercised to ensure that water and foods consumed by HIV/AIDS patients are safe, as contaminated water and foods can cause secondary infections in these patients. [source] Evidence of enhanced bacterial invasion during Diplostomum spathaceum infection in European grayling, Thymallus thymallus (L.)JOURNAL OF FISH DISEASES, Issue 2 2006P Pylkkö Abstract Farmed grayling, Thymallus thymallus (L.), are susceptible to atypical Aeromonas salmonicida (aAS) infections. Interactions between bacteria and parasites were studied using grayling subjected to concomitant exposure to aAS bacteria and the digenean parasite Diplostomum spathaceum. Atypical AS was detected from fish by a combination of bacterial cultivation and polymerase chain reaction techniques. A detection level of 17 aAS cells per 100 mg intestine tissue sample was obtained. Concomitant bacterial exposure did not enhance the severity of grayling eye rupture and nuclear extrusion induced by D. spathaceum, but D. spathaceum invasion into grayling increased the proportion of fish carrying aAS in their heart tissue. However, the number of aAS cells detected in heart tissue was low. Atypical AS did not cause acute disease or mortality during 15 days post-exposure. There was a higher prevalence of aAS in grayling heart samples than in intestinal samples, indicating that the intestine is not favoured by aAS. We suggest that heart tissue would be a good organ from which to isolate aAS when tracing latent carrier fish. We conclude that penetrating diplostomids can enhance bacterial infections in fish and that diplostomids can cause serious eye ruptures in grayling. [source] Use of polymerase chain reaction techniques and sodium dodecyl sulfate-polyacrylamide gel electrophoresis for differentiation of oral Lactobacillus speciesMOLECULAR ORAL MICROBIOLOGY, Issue 2 2006R. Teanpaisan Background/aims:, The genus Lactobacillus has been associated with dental caries in humans, although it is seldom speciated due to lack of simple and nonlaborious identification methods. A considerable heterogeneity among Lactobacillus species has been demonstrated. The purpose of this study was to develop simple methods combining restriction fragment length polymorphism analysis of polymerase chain reaction (PCR)-amplified 16S rRNA (16S rRNA gene PCR-RFLP) and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) for the identification of 13 reference strains of Lactobacillus. Methods:, The 16S rRNA gene sequences were amplified by PCR using universal primers and digestion of PCR products with the restriction endonucleases, HpaII and HaeIII. The 16S rRNA gene PCR-RFLP is reproducible and has been proved to be useful for differentiating Lactobacillus strains to species level. Seventy-seven Lactobacillus isolates from a Thai population were used to show the applicability of the identification test. Results:, PCR-RFLP alone had limitations, because the RFLP patterns of Lactobacillus casei and Lactobacillus rhamnosus and of Lactobacillus acidophilus and Lactobacillus crispatus showed similar patterns; however, these could be differentiated by SDS-PAGE. Of the 77 isolates, 38 were identified as Lactobacillus fermentum, 25 as L. rhamnosus, 5 as Lactobacillus salivarius, 5 as L. casei, 3 as L. acidophilus and 1 as Lactobacillus plantarum. Conclusion:, 16S rRNA gene PCR-RFLP, using HpaII and HaeIII, together with SDS-PAGE protein profiles could be an alternative method for the identification of oral Lactobacillus strains to species level, and may be applicable for large-scale studies on the association of Lactobacillus to dental caries. [source] Recurrent Respiratory Papillomatosis: A Longitudinal Study Comparing Severity Associated With Human Papilloma Viral Types 6 and 11 and Other Risk Factors in a Large Pediatric Population,THE LARYNGOSCOPE, Issue S104 2004Brian J. Wiatrak MD Abstract Objectives/Hypothesis: A database was developed for prospective, longitudinal study of recurrent respiratory papillomatosis (RRP) in a large population of pediatric patients. Data recorded for each patient included epidemiological factors, human papilloma virus (HPV) type, clinical course, staged severity of disease at each surgical intervention, and frequency of surgical intervention. The study hypothesizes that patients with HPV type 11 (HPV-11) and patients younger than 3 years of age at diagnosis are at risk for more aggressive and extensive disease. Study Design: The 10-year prospective epidemiological study used disease staging for each patient with an original scoring system. Severity scores were updated at each surgical procedure. Methods: Parents of children with RRP referred to the authors' hospital completed a detailed epidemiological questionnaire at the initial visit or at the first return visit after the study began. At the first endoscopic debridement after study enrollment, tissue was obtained and submitted for HPV typing using polymerase chain reaction techniques and in situ hybridization. Staging of disease severity was performed in real time at each endoscopic procedure using an RRP scoring system developed by one of the authors (B.J.W.). The frequency of endoscopic operative debridement was recorded for each patient. Information in the database was analyzed to identify statistically significant relationships between extent of disease and/or HPV type, patient age at diagnosis, and selected epidemiological factors. Results: The study may represent the first longitudinal prospective analysis of a large pediatric RRP population. Fifty-eight of the 73 patients in the study underwent HPV typing. Patients infected with HPV-11 were significantly more likely to have higher severity scores, require more frequent surgical intervention, and require adjuvant therapy to control disease progression. In addition, patients with HPV-11 RRP were significantly more likely to develop tracheal disease, to require tracheotomy, and to develop pulmonary disease. Patients receiving a diagnosis of RRP before 3 years of age had significantly higher severity scores, higher frequencies of surgical intervention, and greater likelihood of requiring adjuvant medical therapy. Patients with Medicaid insurance had significantly higher severity scores and required more frequent surgical debridement. Birth by cesarean section appeared to be a significant risk factor for more severe disease and necessity of more frequent surgical intervention. Conclusion: Statistical analysis of the relationships among epidemiological factors, HPV type, and clinical course revealed that patients with HPV-11 and patients younger than 3 years of age at RRP diagnosis are prone to develop more aggressive disease as represented by higher severity scores at endoscopic debridement, more frequent operative debridement procedures per year, a greater requirement for adjuvant therapy, and greater likelihood of tracheal disease with tracheotomy. [source] Timing of Human Insulin-Like Growth Factor-1 Gene Transfer in Reinnervating Laryngeal Muscle,THE LARYNGOSCOPE, Issue 4 2004Hideki Nakagawa MD Abstract Objectives/Hypothesis The authors have designed a rat laryngeal paralysis model to study gene transfer strategies using a muscle-specific expression system to enhance local delivery of human insulin-like growth factor-1 (hIGF-1). In preliminary studies, a nonviral vector containing the ,-actin promoter and human hIGF-1 sequence produced both neurotrophic and myotrophic effects 1 month after single injection of plasmid formulation into paralyzed rat thyroarytenoid muscle in vivo. Based on these findings, it is hypothesized that the effects of hIGF-1 will enhance the results of laryngeal muscle innervation procedures. The timing of gene delivery relative to nerve repair is likely to be important, to optimize the results. Study Design Prospective analysis. Methods The effects of nonviral gene transfer for the delivery of hIGF-1 were evaluated in rats treated immediately following recurrent laryngeal nerve transection and repair and in rats receiving a delayed treatment schedule, 30 days after nerve transection and repair. Gene transfer efficiency was determined using polymerase chain reaction and reverse transcriptase,polymerase chain reaction techniques. Muscle fiber diameter, motor endplate length, and percentage of motor endplates with nerve contact were examined to assess hIGF-1 trophic effects. Results Compared with reinnervated untreated control samples, both early and delayed hIGF-1 transfer resulted in significant increase in muscle fiber diameter. Motor endplate length was significantly decreased and nerve/motor endplate contact was significantly increased following delayed gene transfer, but not after early treatment. Conclusion We infer from results of the study that delayed hIGF-1 gene transfer delivered by a single intramuscular injection will enhance the process of muscle reinnervation. The clinical relevance of these findings supports the future application of gene therapy using nonviral vectors for management of laryngeal paralysis and other peripheral nerve injuries. [source] Prevalence of Herpes Simplex Virus in Malignant Laryngeal Lesions,THE LARYNGOSCOPE, Issue 2 2000Anna M. Pou MD Abstract Objective: To determine the prevalence of herpes simplex virus (HSV) in malignant laryngeal lesions. Study Design: Retrospective review. Materials and Methods: Paraffin-embedded, histologically confirmed specimens containing benign laryngeal lesions, squamous cell carcinoma of the larynx, and squamous cell carcinoma of the oral cavity were identified from archived surgical specimens. Biopsies of normal-appearing oral cavity tissue were also obtained from fresh-frozen cadavers. These tissues were analyzed for the presence of HSV DNA using polymerase chain reaction techniques. Patient charts were reviewed for patient demographics, risk factors, stage, clinical course, treatment, and outcome. Results: HSV was detected in nine laryngeal squamous cell carcinomas (75%) and in none of the benign laryngeal lesions (P = .0001). HSV was also found in three oral cavity squamous cell carcinomas (25%) and in none of the controls (P = .049). Conclusion: HSV is more prevalent in squamous cell carcinoma of the larynx and oral cavity than in their respective control groups, suggesting a role for carcinogenesis. HSV is more prevalent in squamous cell carcinoma of the larynx than of the oral cavity. Further studies to determine the role of HSV as a cocarcinogen and its interrelationship with other environmental factors in laryngeal cancer are warranted. [source] Varicella zoster virus is not a disease-relevant antigen in multiple sclerosis,ANNALS OF NEUROLOGY, Issue 4 2009Mark P. Burgoon PhD Herpesvirions and varicella zoster virus (VZV) DNA were recently reported in all 15 cerebrospinal fluid (CSF) samples from patients with relapsing-remitting multiple sclerosis (MS) obtained within 1 week of exacerbation. Using identical electron microscopic and polymerase chain reaction techniques, including additional primer sets representing different regions of the VZV genome, we found no herpesvirions or VZV DNA in MS CSF or acute MS plaques. Although enzyme-linked immunosorbent assay analysis demonstrated a higher titer of VZV antibody in MS CSF than in inflammatory control samples, recombinant antibodies prepared from clonally expanded MS CSF plasma cells did not bind to VZV. VZV is not a disease-relevant antigen in MS. Ann Neurol 2009;65:474,479 [source] | ||||||||||