Molecular Detection of Adenoviruses in the Sinus Tissues of Patient by Nested-PCR in Shiraz, Southwest Iran

2019 ◽  
Author(s):  
Shahab Mahmoudvand ◽  
Mohammad Kord ◽  
Neda Pirbonyeh ◽  
Afagh Moattari ◽  
Somayeh Shokri ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Molecular Detection ◽  
Viral Infections ◽  
Globin Gene ◽  
Nested Polymerase Chain Reaction ◽  
High Prevalence ◽  
Polymerase Chain ◽  
Southwest Iran

Background and Aims: Rhinosinusitis is an inflammation of the mucous membrane that may be caused by infectious agents such as bacteria, fungi and viruses. Few studies have been carried out on the role of viruses in Rhinosinusitis patients .The aim of this study was the molecular detection of Adenoviruses in sinus tissues by nested polymerase chain reaction (PCR) in Shiraz.Materials and Methods: In the present study, 103 paraffin-embedded biopsy specimens of sinus tissues were subjected to DNA extraction and tested for adenovirus DNA using Nested PCR. The amplification of a β-globin gene by PCR-based method was used to confirm the quality of extracted DNA.Results: A total of 103 samples of sinus tissues were examined. Of these patients, 50 (48.54%) were male and the rest were female (51.46%). The patients’ age ranged between 2 and 82 years and the mean age was 42.15±1.56 years. The adenovirus DNA was detected in 13 of 103 (12.6%) samples.Conclusions: The results of this study showed that Adenoviruses have high prevalence in rhinosinusitis patients. As a results, it is an important to investigate clinical significance of viral infections especially Adeno viruses in these patients.

Epidemiology of Epizootic Lymphangitis of cart horses in Northern Ethiopia Using Conventional diagnostic Methods and Nested Polymerase Chain Reaction

2019 ◽  
Author(s):  
Birhanu Hadush Abera ◽  
Molla Michaelay ◽  
Habtamu Taddele ◽  
Nigus Abebe ◽  
Abrha Tesfay ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Clinical Examination ◽  
Roc Curve ◽  
Nested Pcr ◽  
Control Measure ◽  
Diagnostic Methods ◽  
Northern Ethiopia ◽  
Chain Reaction ◽  
Nested Polymerase Chain Reaction ◽  
Polymerase Chain

Abstract Background: Epizootic lymphangitis (EL), caused by Histoplasma capsulatum variety farciminosum (HCF) is a contagious chronic disease of equines characterized by development of nodular lesions in the lymph nodes, lymphatic vessels and skin. This disease is the most important diseases of equines in Ethiopia causing a significant economic loss, particularly cart pulling equines. Todate there is no sound diagnostic nor control measure implemented in the country. Furthermore, there is a shortage of data on the epidemiology of the disease in different regions of the country including northern Ethiopia. This study was conducted to investigate the epidemiology of EL in northern Ethiopia using the conventional methods and the nested polymerase chain reaction (PCR). Methods: A total of 191 cart-horses were enrolled and used as sources of pus and blood samples. The blood was used for the extraction of the DNA of HCF from buffy coat for nested PCR while the pus samples were cultured on Sabourauds Dextrose Agar for isolation. Statistical Package for Social Sciences (SPSS) version 21 was used for data analysis by applying logistic regression, receiver operating characteristic (ROC) curve and Cohen’s kappa coefficient test. In addition, the level of agreement between the clinical examination and the nested PCR was evaluated. Results: Infection with HCF was confirmed in 44% (84/191) of the horses using nested PCR. Subclinical infection was observed in 18.18% (22/121) of the apparently healthy horses. Considering nested PCR as a gold standard, the sensitivity and specificity of the clinical examination were 74% and 95%, respectively while the area under the ROC curve (AUR) was 0.83 (95% confidence interval, 0.77, 0.896). Moreover, a moderate (k=0.675) agreement was observed between the nested PCR and clinical examination.Conclusions: The findings of the present study showed the wide spread occurrence of EL in northern Ethiopia and the advantage of the nested PCR in detecting of the infection of HCF even before the clinical symptoms are apparent.

scholarly journals In SituReverse Transcriptase-Nested Polymerase Chain Reaction to Identify Intracellular Nucleic Acids without the Necessity of Dnase Pretreatment and Hybridisation

2001 ◽  
Vol 22 (3) ◽  
pp. 151-158 ◽  
Author(s):  
Mario Menschikowski ◽  
Margot Vogel ◽  
Rolf Eckey ◽  
Gerd Dinnebier ◽  
Werner Jaross
Keyword(s):  
Polymerase Chain Reaction ◽  
Nucleic Acids ◽  
Nested Pcr ◽  
In Situ Hybridisation ◽  
Target Sequence ◽  
Chain Reaction ◽  
Multiple Control ◽  
Nested Polymerase Chain Reaction ◽  
Polymerase Chain

In the present study a protocol of in situ reverse transcriptase‐nested polymerase chain reaction (in situ RT‐nested PCR) was examined based on the following modifications. (i) To exclude false positive signals caused by “DNA repair mechanisms” and “endogenous priming”, a two‐step PCR was applied after reverse transcription. The first step was performed in the presence of extrinsic primers and unlabeled nucleotides with a maximum of PCR cycles possible without destroying the cell morphology. The second step consisted of only one annealing/elongation reaction, the target sequence marked by addition of digoxigenin‐labeled nucleotides and intrinsic primers. (ii) In order to prevent amplifications of genomic DNA nested primer pairs were applied crossing intron sequences. (iii) To minimize the diffusion of PCR products in cells, the extrinsic primers were extended with complementary 5′‐tails. This approach results in the generation of high molecular weight concatamers during PCR cycles. By applying this protocol, immunostainings specific for phospholipase A2 of type IIA mRNA were exclusively detectable in the cytoplasm of HepG2 hepatoma cells, which were used as a model system, whereas the nuclei were unstained. Multiple control experiments yielded completely negative results. These data suggest that the in situ RT‐nested PCR, which in comparison to the method of in situ RT‐PCR‐in situ‐hybridisation is simpler and less time‐consuming, can be used as an alternative approach to identify intracellular nucleic acids.

Application of Nested Polymerase Chain Reaction to Detection of Salmonella in Poultry Environment

2002 ◽  
Vol 65 (8) ◽  
pp. 1227-1232 ◽  
Author(s):  
TONGRUI LIU ◽  
KAREN LILJEBJELKE ◽  
ELIZABETH BARTLETT ◽  
CHARLES HOFACRE ◽  
SUSAN SANCHEZ ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Nested Pcr ◽  
Virulence Gene ◽  
Pcr Primers ◽  
Processing Plant ◽  
Chain Reaction ◽  
Chi Square ◽  
Nested Polymerase Chain Reaction ◽  
Secondary Enrichment ◽  
Polymerase Chain

Isolation of Salmonella from environmental and processing-plant poultry samples requires the sampling of large numbers of areas within the poultry house or plant. Subsequently, the required number of samples necessitates a large volume of work for a microbiology laboratory, especially when the protocol requires the inclusion of a delayed secondary enrichment for the isolation of Salmonella. This study examined the use of the polymerase chain reaction (PCR) to identify those secondary enrichments containing Salmonella. The unique Salmonella virulence gene invA was chosen as the target for the development of a nested PCR because of its uniform distribution among Salmonella serotypes. The use of nested PCR primers increased the sensitivity of detection 100-fold, resulting in the detection of as few as four cells. There was a strong, statistically significant positive correlation between PCR and culture results as determined by chi-square (P < 0.001) and kappa (k = 0.915; excellent agreement) tests. Using PCR to screen primary enrichments for presumptive Salmonella contamination, we improved our efficiency at isolating Salmonella upon secondary enrichment by 20%, and no false negatives were observed. This method will not only validate the use of secondary enrichment procedures but also reduce costs and manpower required for the surveillance of Salmonella.

scholarly journals Occurrence of a 16SrIV Group Phytoplasma not Previously Associated with Palm Species in Yucatan, Mexico

Plant Disease ◽  
2011 ◽  
Vol 95 (3) ◽  
pp. 256-262 ◽  
Author(s):  
Roberto Vázquez-Euán ◽  
Nigel Harrison ◽  
María Narvaez ◽  
Carlos Oropeza
Keyword(s):  
Cocos Nucifera ◽  
Restriction Enzymes ◽  
Nested Polymerase Chain Reaction ◽  
Palm Trees ◽  
Palm Species ◽  
Leaf Yellowing ◽  
Sabal Mexicana ◽  
Polymerase Chain ◽  
First Time

The occurrence of 16SrIV group phytoplasmas in palm species Sabal mexicana and Pseudophoenix sargentii is reported here for the first time. Palm trees showed leaf decay and leaf yellowing syndromes, respectively. An amplification product (1.4 kb) was obtained in symptomatic S. mexicana (18 of 21) and symptomatic P. sargentii (1 of 1) palm trees sampled in different locations in Yucatan State, Mexico; five of the positive S. mexicana and the positive P. sargentii trees died. The identity of the phytoplasmas from these species was determined by restriction fragment length polymorphism profiling with restriction enzymes AluI and HinfI, showing there could be two phytoplasma strains of the 16SrIV group. In one S. mexicana palm, the profile was the same as observed with these enzymes for phytoplasmas of 16SrIV-A subgroup, previously associated with Cocos nucifera palm trees and, in the rest of the trees, including the P. sargentii palm, the profile was for phytoplasmas of the 16SrIV-D subgroup. These identities were supported by analyses of the amplicons obtained by nested polymerase chain reaction by nucleotide-nucleotide BLAST analysis. Geographical distribution of the association S. mexicana/16SrIV group phytoplasmas was found widely dispersed in Yucatan State. A potential role of S. mexicana palm trees as a permanent source of phytoplasma inoculum is suggested. In addition to P. sargentii, other palm species (Thrinax radiata and C. nucifera) coexisting with S. mexicana trees were also sampled and analyzed.

Nested PCR protocol for the rapid detection of Escherichia coli in potable water

1996 ◽  
Vol 42 (8) ◽  
pp. 862-866 ◽  
Author(s):  
David Juck ◽  
Jordan Ingram ◽  
Michèle Prévost ◽  
Josée Coallier ◽  
Charles Greer
Keyword(s):  
Escherichia Coli ◽  
Nested Pcr ◽  
Potable Water ◽  
Test Organism ◽  
Bacterial Cells ◽  
Nested Polymerase Chain Reaction ◽  
Fecal Indicator ◽  
E Coli ◽  
Pcr Products ◽  
Polymerase Chain

A rapid and sensitive method for the detection of low levels of bacteria in potable water was developed. The fecal indicator bacterium Escherichia coli was used as the test organism in a filtration concentration–nested polymerase chain reaction (PCR) protocol, combined with ethidium bromide visualization of PCR products. Two sets of primers were designed from the E. coli specific β-glucuronidase gene (uidA), the primary pair producing a 486-bp fragment that was used as template for the nested primer pair delineating a 186-bp fragment. This protocol can detect 1–10 bacterial cells/50 mL water sample within 6–8 h, in contrast to traditional culturing or Southern hybridization methods which require 2–3 days for results.Key words: nested PCR, sensitive, detection, potable water.

scholarly journals High prevalence of campylobacter excretors among Liberian children related to environmental conditions

1988 ◽  
Vol 100 (2) ◽  
pp. 227-237 ◽  
Author(s):  
Kåre Mølbak ◽  
Niels Højlyng ◽  
Knud Gaarslev
Keyword(s):  
Environmental Contamination ◽  
Environmental Conditions ◽  
High Intensity ◽  
Bacterial Pathogen ◽  
Urban Slum ◽  
Pathogenic Role ◽  
High Prevalence ◽  
Very High

SUMMARYCampylobacter was the bacterial pathogen most prevalent in 850 children, aged 6–59 months, examined in a house-to-house diarrhoea survey in two Liberian communities. 44·9% of the children from an urban slum and 28·4% from a rural area were excretors. Since the prevalence of diarrhoea was very high and consequently many convalescent carriers were found, it was not possible to evaluate the pathogenic role of campylobacter.The excretor rate increased with ago and was significantly correlated to the uso of supplementary feeding, inversely correlated to the quality of the water supply, and also associated with helminthic infestation. Results from re-examination of 172 children suggested a high intensity of transmission.The findings all indicate the existence of a heavy environmental contamination with campylobacter, probably of both human and animal faecal origin.

scholarly journals Detection of NaturalToxoplasma gondiiInfection in Chicken in Thika Region of Kenya Using Nested Polymerase Chain Reaction

2016 ◽  
Vol 2016 ◽  
pp. 1-5 ◽  
Author(s):  
John Mokua Mose ◽  
John Maina Kagira ◽  
Simon Muturi Karanja ◽  
Maina Ngotho ◽  
David Muchina Kamau ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Toxoplasma Gondii ◽  
Human Beings ◽  
Chain Reaction ◽  
Nested Polymerase Chain Reaction ◽  
Free Range ◽  
Age Dependent ◽  
Polymerase Chain ◽  
The Difference

The detection ofToxoplasma gondiiin free-range chickens is a good indicator of possible risk to human beings. The aim of this study was to investigate the occurrence ofT. gondiiin free-range chicken using polymerase chain reaction (PCR). Brain samples from 105 free-range chickens from three administrative areas in Thika region, Kenya, were collected, DNA-extracted, and analyzed using PCR to detect presence ofT. gondii. The overall prevalence ofT. gondiiin all the three areas was 79.0% (95% CI: 70.0–86.4%) and the prevalence across the three areas was not significantly different (P=0.5088;χ2=1.354). Female chickens had higher (79.4%) prevalence than males (78.6%), although the difference was not significant (P=0.922,χ2= 0.01). However, chickens that were more than 2 years old had significantly (P=0.003;χ2= 11.87) higher prevalence compared to younger ones. The study indicates that there was a high occurrence ofT. gondiiinfection in free-range chickens from Thika region and that the infection rate is age dependent. Further studies should be carried out to determine the possible role of roaming chickens in the epidemiology of the disease among humans in the area.

scholarly journals The role of the dentist and orthodontist in recognizing oro-facial manifestations of acromegaly: a questionnaire-based study

Pituitary ◽  
2021 ◽  
Author(s):  
Giorgia Preo ◽  
Alberto De Stefani ◽  
Francesca Dassie ◽  
Alexandra Wennberg ◽  
Roberto Vettor ◽  
...  
Keyword(s):  
Quality Of Life ◽  
Early Recognition ◽  
Sella Turcica ◽  
Mandibular Prognathism ◽  
Oral Manifestation ◽  
Number Of Patients ◽  
High Prevalence ◽  
Strategic Role

Abstract Purpose Oro-facial manifestations of acromegaly are among the earliest signs of the disease and are reported by a significant number of patients at diagnosis. Despite this high prevalence of acromegaly oral manifestation, dentists do not play a pivotal role in acromegaly identification and diagnosis. The aim of our study was to evaluate the ability of dentists and orthodontists in the early recognition of the oro-facial manifestations of acromegaly. Methods A telematic questionnaire was administered to dentists and orthodontists. The questionnaire included photos with facial and oral-dental details and lateral teleradiography of acromegaly patients (ACRO). Results The study included 426 participants: 220 dentists and 206 orthodontists. Upon reviewing the photos, dentists most often observed mandibular prognathism and lips projection, while orthodontists also reported the impairment of relative soft tissue. Orthodontists, who usually use photos to document patients’ oral-facial characteristics, paid more attention to oral-facial impairment than dentists. During dental assessment, 90% of the participants usually evaluated tongue size and appearance, diastemas presence, and signs of sleep impairment (mainly orthodontists). Orthodontists were also more able to identify sella turcica enlargement at teleradiography. A total of 10.8% of the participants had ACRO as patients and 11.3% referred at least one patient for acromegaly suspicion. Conclusion The study highlighted dentists’ strategic role in identifying ACRO. Increasing dentists’ awareness about acromegaly clinical issues may improve early diagnosis, potentially resulting in an increased quality of life and decreased mortality among ACRO.

scholarly journals Osteoarticular tuberculosis – diagnostic solutions in a disease endemic region

2009 ◽  
Vol 3 (07) ◽  
pp. 511-516 ◽  
Author(s):  
Vikas Agashe ◽  
Shubhada Shenai ◽  
Ganesh Mohrir ◽  
Minal Deshmukh ◽  
Anita Bhaduri ◽  
...  
Keyword(s):  
Nested Pcr ◽  
Bone Diseases ◽  
Drug Resistant ◽  
Endemic Region ◽  
Mycobacterial Culture ◽  
Osteoarticular Tuberculosis ◽  
Private And Public ◽  
Polymerase Chain ◽  
Negative Controls

Background: We conducted a study of osteoarticular tuberculosis in patients from private and public settings in a disease endemic area. Our objective was to assess the role of mycobacterial culture and polymerase chain reaction (PCR) in the diagnosis of osteoarticular tuberculosis (TB) in settings where only clinical and imaging diagnosis form the basis for treatment. Methodology: Ninety-three consecutive specimens collected from clinically suspected patients of osteoarticular TB were screened for bacterial culture, mycobacterial culture and in-house nested PCR. In addition, specimens were examined by imaging and histopathology. Ten specimens collected from patients suffering from other bone diseases were included as negative controls. Results: Of the 93 clinically suspected TB patients, mycobacterial culture was positive for Mycobacterium tuberculosis (MTB) in 47 (51%) patients who were confirmed as definite TB cases. Of the remaining patients, 16 (17%) were diagnosed as probable, 19 (20%) as possible, and 11 (12%) as only clinically suspected TB cases. In-house nested PCR was positive in 65 (70%) cases. Fifteen patients were resistant to one or more anti-tuberculous drugs; twelve patients were multi-drug resistant, two of whom were extensively drug resistant. Conclusion: Mycobacterial cultures using liquid media with susceptibility should form the backbone of management of osteoarticular TB. Nested PCR enhances the sensitivity if performed in addition to culture.

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