Development and evaluation of the polymerase chain reaction (PCR) method for diagnosis of spring viremia of carp virus (SVCV)

Aquaculture has been expanded rapidly to become a major commercial and food-producing sector worldwide in recent decade. In parallel, viral diseases rapidly spread among farms causing enormous economic losses. The accurate detection of pathogens at early stages of infection is a key point for disease control in aquaculture. Spring Viraemia of Carp Virus (SVCV) is a very severe pathogen of carp fishes in different parts of the world and is categorized as a reportable listed disease in the annual published list of World Organization for animal Health (OIE). The objective of this study was to develop and evaluate RT- PCR test for detecting SVC virus and also the sensitivity and specificity of this test. A semi nested RT- PCR was designed using combination of three primers: two external (SVCF , SVCR) and one internal (SVCS) primers which based on conserved region of G gen. The specificity of designed primers (only external ones) by examination on Viral Hemorrhagic Septicemia Virus (VHSV) and Infectious Hematopoietic Necrosis Virus (IHNV) was confirmed. For optimizing of the PCR test, primer concentration, primer annealing temperature, cycle number and Mgcl2 concentration were surveyed. Also for validity test, prevention of false negative and Assurance of its accuracy, a competitive internal control (mimic) designed and its suitable concentration was defined. Evaluation of the sensitivity of designed test were conducted first by comparing the different commercially available RNA isolation guidelines, two guidelines: isotiocyanate phenol–chloroform based protocols (RNX–Plus Iran, Iq2000 kit Taiwan ) and two column based protocols (Cinna pure RNA Iran , high pure viral RNA kit, Roche Germany ). The results indicated that the column based protocols (Roche method and Cinna pure), yield 36.77 ng/μl and 16/47 ng/μl RNA concentration respectively, which were significantly higher than other protocols(P<0.05). Then for evaluation of extracted RNA sensitivity, Serial dilution of SVCV strain 56.70 grown in EPC (1.9×105 TCID50/ml) was examined To compare sensitivity. Extracted RNA from serial dilution with stone's primers and commercial IQ-2000 kit were examined simultaneously. The result indicated that designed semi- nested RT- PCR was able to recognize SVC virus to 10-4 dilution and stone's primer recognize to 10-3 dilution whereas Iq-2000 commercial kit did not recognized in any dilution. In high virus titer in designed test two DNA band (462 bp and 266 bp) produced, and by decreasing virus titer 462 bp was omitted. In low virus titer or lack of virus, just DNA band (mimic) 729 bp can propagate. After designing and optimizing PCR test, a total of 400 suspected cultured Cyprinus carpio with high mortality from 4 aquaculture zone of Khuzestan province were collected and tested for SVCV during 2012- 2013 using developed PCR method and IQ- 2000. The results indicated that SVC virus was not observed in samples using both methods.

Field study and numerical simulation of developing red tide in the northern Strait of Hormuz

The catastrophic event of red tide has happened in the Strait of Hormuz, the Persian Gulf and Gulf of Oman from late summer 2008 to spring 2009. With its devastating effects, the phenomenon shocked all the countries located in the margin of the Persian Gulf and the Gulf of Oman and caused considerable losses to fishery industries, tourism, and tourist and trade economy of the region. In the maritime cruise carried out by the Persian Gulf and Gulf of Oman Ecological Research Institute, field data, including temperature, salinity, chlorophyll-a, dissolved oxygen and algal density were obtained for this research. Satellite information was received from MODIS and MERIS and SeaWiFS sensors. Temperature and surface chlorophyll images were obtained and compared with the field data and data of PROBE model. The results obtained from the present research indicated that with the occurrence of harmful algal blooms (HAB), the Chlorophyll-a and the dissolved oxygen contents increased in the surface water. Maximum algal density was seen in the northern coasts of the Strait of Hormuz. Less concentration of algal density was detected in deep and surface offshore water. Our results show that the occurred algal bloom was the result of seawater temperature drop, water circulation and the adverse environmental pollutions caused by industrial and urban sewages entering the coastal waters in this region of the Persian Gulf ,This red tide phenomenon was started in the Strait of Hormuz and eventually covered about 140,000 km2 of the Persian Gulf and total area of Strait of Hormuz and it survived for 10 months which is a record amongst the occurred algal blooms across the world. Temperature and chlorophyll satellite images were proportionate to the measured values obtained by the field method. This indicates that satellite measurements have acceptable precisions and they can be used in sea monitoring and modeling.