Establishment of rapid detection method and surveillance of budgerigar fledgling disease virus using a TaqMan Real-Time PCR

Bacillus anthracis, the causative agent of anthrax, has the capacity to form highly resilient spores as part of its life cycle. The potential for the dissemination of these spores using food as a vehicle is a huge public health concern and, hence, requires the development of a foodborne bioterrorism response approach. In this work, we address a critical gap in food biodefense by presenting a novel, combined, sequential method involving the use of real-time PCR and pyrosequencing for the rapid, specific detection of B. anthracis spores in three food matrices: milk, apple juice, and bottled water. The food samples were experimentally inoculated with 40 CFU ml−1, and DNA was extracted from the spores and analyzed after immunomagnetic separation. Applying the combination of multiplex real-time PCR and pyrosequencing, we successfully detected the presence of targets on both of the virulence plasmids and the chromosome. The results showed that DNA amplicons generated from a five-target multiplexed real-time PCR detection using biotin-labeled primers can be used for single-plex pyrosequencing detection. The combined use of multiplexed real-time PCR and pyrosequencing is a novel, rapid detection method for B. anthracis from food and provides a tool for accurate, quantitative identification with potential biodefense applications.

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Rapid detection of foot-and-mouth disease virus using a field-portable nucleic acid extraction and real-time PCR amplification platform

The Veterinary Journal ◽

10.1016/j.tvjl.2011.10.017 ◽

2012 ◽

Vol 193 (1) ◽

pp. 67-72 ◽

Cited By ~ 24

Author(s):

Mikidache Madi ◽

Andrea Hamilton ◽

David Squirrell ◽

Valerie Mioulet ◽

Phillip Evans ◽

...

Keyword(s):

Nucleic Acid ◽

Real Time ◽

Disease Virus ◽

Real Time Pcr ◽

Rapid Detection ◽

Pcr Amplification ◽

Foot And Mouth Disease ◽

Acid Extraction ◽

Nucleic Acid Extraction ◽

Mouth Disease Virus

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Rapid detection of Chlamydia/Chlamydophila group in samples collected from swine herds with and without reproductive disorders

Polish Journal of Veterinary Sciences ◽

10.2478/pjvs-2014-0052 ◽

2014 ◽

Vol 17 (2) ◽

pp. 367-369 ◽

Cited By ~ 3

Author(s):

K. Rypula ◽

A. Kumala ◽

P. Lis ◽

K. Niemczuk ◽

K. Płoneczka-Janeczko ◽

...

Keyword(s):

Real Time ◽

Real Time Pcr ◽

Rapid Detection ◽

Complement Fixation Test ◽

Complement Fixation ◽

Fixation Test ◽

Reproductive Disorders ◽

Serum Samples ◽

Swine Herds

Abstract The study was carried out in seven reproductive herds of pigs. In three of them reproductive disorders were observed. Three herds consisted of 10-50 and four consisted of 120-500 adult sows and they were called small and medium, respectively. Fifty-seven adult sows were randomly selected from herds. Serum samples were tested using the complement fixation test and swabs from both eyes and from the vaginal vestibule were examined using real-time PCR. All serum samples were negative. Infected sows were present in each of the study herds. In total, there were 28 positive samples (53%, 28/48) in real-time PCR in sows with reproductive disorders and 35 (53%, 35/66) in sows selected from herds without problems in reproduction. One isolate proved to be Chlamydophila pecorum, whereas all the remaining were Chamydia suis

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Faculty Opinions recommendation of Rapid detection of eight causative pathogens for the diagnosis of bacterial meningitis by real-time PCR.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.713998097.789552812 ◽

2012 ◽

Author(s):

Sanjay Tyagi

Keyword(s):

Bacterial Meningitis ◽

Real Time ◽

Real Time Pcr ◽

Rapid Detection

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Isolation and identification of Marek’s disease virus (MDV) from feather follicle epithelium and internal organs of diseased chickens in Dakahlia Governorate, Egypt

Mansoura Veterinary Medical Journal ◽

10.35943/mvmj.2019.22.102 ◽

2019 ◽

Vol 20 (2) ◽

pp. 6-11

Author(s):

Aly El-Kenawy ◽

Mohamed El-Tholoth ◽

Emad A

Keyword(s):

Real Time ◽

Disease Virus ◽

Real Time Pcr ◽

Marek's Disease Virus ◽

Marek's Disease ◽

Marek’S Disease Virus ◽

Marek’S Disease ◽

Field Samples ◽

Feather Follicle ◽

Positive Results

In the present study, a total of 16 samples including feather follicle epithelium, ovary, spleen and kidney (4 samples for each organ) were collected from diseased chicken flocks suspected to be infected with Marek’s disease virus (MDV) at Dakahlia Governorate, Egypt during the period from October 2016 to October 2017. Each sample was pooled randomly from three to five birds (90 to 360 days old). The isolation of the suspected virus from the collected samples was carried out via chorioallantoic membranes (CAMs) of 12 days old embryonated chicken eggs (ECEs). Three egg passages were carried out for each sample. Hyperimmune serum was prepared against standard MDV. MDV in both field and egg passaged samples (after 3rd passage) was identified by agar gel precipitation test (AGPT) and indirect fluorescence antibody test (IFAT). Molecular identification of virus was carried out by conventional polymerase chain reaction (PCR) and real- time PCR in four selected samples. The results revealed that 14 samples (87.5%) including 4 (100%) samples from feather follicle epithelium, ovary and kidney and 2 (50%) samples from spleen, showed positive results in virus isolation after 3rd passage. The positive results percentage by AGPT for field samples were 50% (8 out of 16 samples), while after the 3rd passage in ECEs were 37.5% (6 out of 16 samples) and the positive results percentage by IFAT for field samples were 62.5% (10 out of 16 samples), while after the 3rd passage in ECEs were 81.25 % (13 out of 16 samples). Viral nucleic acid was detected in all selected samples by conventional and real- time PCR. The results indicate that feather follicle epithelium is the best organ for MDV detection. IFAT is superior over AGPT in virus detection. Conventional and real - time PCR could be efficiently used for molecular detection of the virus.

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