Volume 24, Issue 5 (Iranian South Medical Journal 2021)
Iran South Med J 2021, 24(5): 469-480 |
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Moein H, Kafilzadeh F, Miri M R, Khalilpazir M. Design and Evaluation of a PCR Method for Detecting White Spot Syndrome Virus in Shrimp. Iran South Med J 2021; 24 (5) :469-480
URL: http://ismj.bpums.ac.ir/article-1-1516-en.html
URL: http://ismj.bpums.ac.ir/article-1-1516-en.html
1- Department of Microbiology, Jahrom Branch, Islamic Azad University, Jahrom, Iran
2- Department of Microbiology, Jahrom Branch, Islamic Azad University, Jahrom, Iran ,f.kafilzadeh@gmail.com
3- The Persian Gulf Marine Biotechnology Research Center, The Persian Gulf Biomedical Sciences Research Insti-tute, Bushehr University of Medical Sciences, Bushehr, Iran
4- Iranian Shrimp Research Center,Iranian Fisheries Science Research Institute, Agricultural Research, Education & Extention Organization (AREEO), Bushehr, Iran
2- Department of Microbiology, Jahrom Branch, Islamic Azad University, Jahrom, Iran ,
3- The Persian Gulf Marine Biotechnology Research Center, The Persian Gulf Biomedical Sciences Research Insti-tute, Bushehr University of Medical Sciences, Bushehr, Iran
4- Iranian Shrimp Research Center,Iranian Fisheries Science Research Institute, Agricultural Research, Education & Extention Organization (AREEO), Bushehr, Iran
Abstract: (2835 Views)
Background: White spot syndrome virus (wssv) is the causing agent for white spot disease in shrimp and many crustaceans. This disease is highly contagious and can cause death within 3–10 days under normal culture conditions. Therefore, early diagnosis of the virus is a necessity.
Materials and Methods: Primers were designed for three regions of the virus genome and one region of the shrimp genome so that they could function together in a reaction. DNA was extracted from 40 samples of shrimp suspected of white spot disease. Primers were then optimized individually to detect white spot virus and after selecting the most suitable one, the virus was detected simultaneously by two pairs of primers. Shrimp genome replication primers were also used as internal control.
Results: Among the designed primers, three pairs of primers were selected that amplified one fragment of the shrimp genome and two fragments of the virus genome. Of the 40 samples examined, 28 samples were positive (infected with the virus) and 12 samples were negative, which completely matched the results obtained with the reference kit.
Conclusion: To detect white spot virus, examination of two regions of the virus genome is sufficient and reduces the possibility of false negatives. It is also effective to use the shrimp genome to control DNA extraction and PCR steps. The single-step method (PCR) is preferable to the two-step method (Nested PCR) due to its reduced probability of contamination and ease of use.
Materials and Methods: Primers were designed for three regions of the virus genome and one region of the shrimp genome so that they could function together in a reaction. DNA was extracted from 40 samples of shrimp suspected of white spot disease. Primers were then optimized individually to detect white spot virus and after selecting the most suitable one, the virus was detected simultaneously by two pairs of primers. Shrimp genome replication primers were also used as internal control.
Results: Among the designed primers, three pairs of primers were selected that amplified one fragment of the shrimp genome and two fragments of the virus genome. Of the 40 samples examined, 28 samples were positive (infected with the virus) and 12 samples were negative, which completely matched the results obtained with the reference kit.
Conclusion: To detect white spot virus, examination of two regions of the virus genome is sufficient and reduces the possibility of false negatives. It is also effective to use the shrimp genome to control DNA extraction and PCR steps. The single-step method (PCR) is preferable to the two-step method (Nested PCR) due to its reduced probability of contamination and ease of use.
Type of Study: Original |
Subject:
Microbiology and Immunology
Received: 2021/11/28 | Accepted: 2021/11/28 | Published: 2021/11/28
Received: 2021/11/28 | Accepted: 2021/11/28 | Published: 2021/11/28
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