The use of reverse transcription-polymerase chain reaction (RT-PCR) for the rapid detection and identification of dengue virus in an endemic region: a validation study

Author(s):  
Sérgio Oliveira De Paula ◽  
Roberto J. Pires^Neto ◽  
Joseane A.C. Tocantins Corrêa ◽  
Silvia R. Assumpção ◽  
Márcia L.S. Costa ◽  
...  
2005 ◽  
Vol 95 (2) ◽  
pp. 166-171 ◽  
Author(s):  
Hiroyuki Uga ◽  
Shinya Tsuda

A one-step reverse transcription-polymerase chain reaction (RT-PCR) method has been developed for the simultaneous detection and identification of multiple tospoviruses that infect plants. The RT-PCR system is composed of six primers in a single tube: a universal degenerate primer and five virus species-specific primers. Amplifications resulted in an 848-bp PCR product for Watermelon silver mottle virus, 709-bp for Tomato spotted wilt virus, 589-bp for Impatiens necrotic spot virus, 511-bp for Melon yellow spot virus, and a 459-bp amplicon for Iris yellow spot virus. This system enables the simultaneous detection of at least three types of tospovirus infections, in addition to their species identities, from five possible tospoviruses studied, on the basis of their S RNA combinations. This multiplex RT-PCR system was applied to the detection of tospovirus in ornamental crops cultivated in fields and shows potential for epidemiological studies.


2016 ◽  
Vol 4 (2) ◽  
Author(s):  
Grace Trovancia ◽  
Angle Sorisi ◽  
Josef S.B. Tuda

Abstract: Dengue hemorrhagic fever is an acute disease with clinical manifestations of hemorrhage caused by dengue virus infection. Manado is endemic dengue. Dengue virus has the ability to maintain its existence in nature through horizontal and vertical transmission. There are several ways to detect the dengue virus by Reverse Transcription-Polymerase Chain Reaction (RT-PCR) and immunohistochemistry Streptavidin Biotin Peroxidase Complex (ISBPC). This research aims to determine the wild Aedes aegypti population in Manado and to detect dengue virus in wild mosquito Aedes aegypti by ISBPC methods. This was a descriptive survey study with a cross sectional design to describe the transmission of dengue virus in wild mosquito Aedes aegypti in the city of Manado. The results showed that there were 5 wild Aedes aegypti mosquitoes positive for dengue virus, and 36 wild Aedes aegypti mosquitoes negative containing dengue virus. Conclusion: Of the 41 samples immunohistochemistry tested, 5 samples showed dengue virus transmission in wild mosquito Aedes aegypti in Manado which is a positive possibility of horizontal transmission.Keywords: detection of dengue virus, transmission, wild Aedes aegypti, Manado. Abstrak: Demam berdarah dengue adalah suatu penyakit akut dengan manifestasi klinis perdarahan yang disebabkan oleh infeksi virus dengue. Manado merupakan daerah endemis demam berdarah. Virus dengue memiliki kemampuan untuk mempertahankan keberadaannya di alam melalui transmisi horizontal dan vertikal. Ada beberapa cara untuk mendeteksi virus dengue yaitu Reverse Transcription-Polymerase Chain Reaction (RT-PCR) dan imunohistokimia Streptavidin Biotin Peroxidase Complex (SBPC). Penelitian ini bertujuan untuk mengetahui populasi nyamuk wild Aedes aegypti di Kota Manado dan mendeteksi virus dengue pada nyamuk wild Aedes aegypti dewasa menggunakan metode imunohistokimia streptavidin biotin peroxidase complex (ISBPC). Jenis penelitian ialah survei deskriptif dengan desain potong lintang untuk mengetahui gambaran transmisi virus dengue pada nyamuk wild Aedes aegypti betina di Kota Manado. Hasil pene;itian mendapatkan 5 nyamuk wild Aedes aegypti positif mengandung virus dengue, dan 36 nyamuk wild Aedes aegypti negatif mengandung virus dengue. Simpulan: Berdasarkan hasil penelitian dapat disimpulkan bahwa dari 41 sampel yang telah diuji imunohistokimia, 5 sampel gambaran transmisi virus dengue pada nyamuk wild Aedes aegypti betina di Kota Manado yang kemungkinan transmisi horizontal adalah positif. Kata kunci: deteksi virus dengue, transmisi, wild Aedes aegypti, Manado.


Author(s):  
A Kadir ◽  
S Ahmed

Foot and mouth disease (FMD) is a major constraint for livestock in Bangladesh; as outbreak of FMD remains uncontrolled despite vaccination. Accurate and Rapid detection of FMD virus with its serotype in field samples is indispensible. So, molecular detection of FMDV was adopted using RT-PCR (reverse transcription- polymerase chain reaction) and mRT-PCR (multiplex reverse transcription-polymerase chain reaction). Ten (10) FMD suspected clinical samples from cattle of two different outbreak areas of Sylhet district of Bangladesh was collected. One set of universal primer (P32:P33) was used in RT-PCR for the detection of FMD virus regardless of their serotypes and a cocktail of primer mix (P38:P40:P74-77:P110) was used in mRT-PCR intending the identification of the serotypes A, O, C and Asia 1. Using universal primer sets 90% of the samples generated amplicon of expected size, indicating the samples containing FMD virus. By mRT-PCR, two serotypes, ‘O’ and Asia 1 were successfully, whereas type C and A were absent in this study. Out of the 9 viruses; 7 was identified as serotype ‘O’ and 2 were identified as Asia-1. Our study indicates that FMDV serotype ‘O’ and Asia-1 was circulating in the two upazilas (Sub-district) of Sylhet district during the study period. Our study also endorses that, RT-PCR and mRT-PCR can successfully be used for a dependable and rapid detection of FMD. However, presence and detection of ‘O’ and Asia-1 serotype of FMDV through this study and serotype A by other researchers emphasizes the critical need for use of trivalent vaccine in the field.International Journal of Natural Sciences (2014), 4(1) 28-34


2006 ◽  
Vol 175 (4S) ◽  
pp. 485-486
Author(s):  
Sabarinath B. Nair ◽  
Christodoulos Pipinikas ◽  
Roger Kirby ◽  
Nick Carter ◽  
Christiane Fenske

2003 ◽  
Vol 15 (2) ◽  
pp. 99 ◽  
Author(s):  
Paisan Tienthai ◽  
Naoko Kimura ◽  
Paraskevi Heldin ◽  
Eimei Sato ◽  
Heriberto Rodriguez-Martinez

Hyaluronan (HA) has been related to fertilization and embryo development in the pig. Furthermore, HA is present in pig oviduct fluid and the lining epithelium, particularly of the pre-ovulatory sperm reservoir. Because the mechanisms that regulate HA synthesis have not yet been clarified, semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) was conducted to assess the expression of mRNAs of two HA-synthesizing enzymes (has2 and has3) in the oviduct epithelium (uterotubal junction, isthmus, ampullary–isthmic junction and ampulla segments) of non-inseminated (control) and inseminated (treatment) sows at pre-, peri- and post-ovulatory oestrus. Only has3 mRNA was detected; it was present in all tubal segments of both control and treatment samples. The level of has3 expression did not vary significantly between non-inseminated and inseminated specimens, but there was a tendency (NS) for increased mean values during the peri- and post-ovulatory stages compared with pre-ovulation. It is concluded that has3 is expressed by the porcine endosalpinx epithelium and the levels of expression do not vary during the critical periods of sperm transport and fertilization, despite fluctuating levels of HA in the tubal fluid at corresponding periods.


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