A primer and kit for early detection of intestinal microsporidia in grouper

Abstract

The invention discloses a method for early detection of grouper intestinal tract microsporidia and application thereof. The method includes the following steps that 1, a fore intestine of grouper is selected, and intestine content is scrapped; 2, genomic DNA of the fore intestine content of grouper is extracted; 3, with the extracted genomic DNA as a template, a specific primer is adopted to conduct PCR amplification; 4, a PCR amplification product in the step 3 is taken and subjected to electrophoretic analysis, a result is judged according to the size of the amplification product, if 450 bp stripes can be amplified through specific amplification, it is proved that the sample is infected by the grouper intestinal tract microsporidia. The problem that diagnosis is difficult as a grouper intestinal tract microsporidia individual is small is solved, programming and standardization of diagnosis are achieved, detection specificity and sensitivity are high, and the method can be applied to diagnosis of grouper intestinal tract microsporidia disease, applied to investigation of the molecular epidemiology and used for assessing whether aquaculture water and bait are suitable for grouper aquaculture or not.

Description

technical field [0001] The invention belongs to the field of animal molecular biology, and in particular relates to a primer and a kit for early detection of grouper intestinal microsporidia. Background technique [0002] Grouper is a warm-water reef fish widely distributed in the tropical and subtropical waters of the Pacific Ocean and the Indian Ocean. (Epinephelinae), is a famous and precious marine fish in the world. It is the main target of mariculture industry in southern China, Guangdong, Hainan, Fujian, Guangxi, etc., with huge economic value (Lin Haoran 2012). However, with the popularization of high-density culture of grouper and cross-contamination of cultured waters, diseases have become one of the main factors restricting the healthy development of grouper aquaculture, such as the nerve necrosis virus in the seedling stage (Liu Xiaodan, 2014) . Since 2011, a new disease called "white stool slimming" has appeared in the main production areas of Hainan grouper, ...

AI Technical Summary

Problems solved by technology

The initial symptom is to stop feeding and pull white feces (commonly known as white feces), then the fish body is obviously thin, the skull and abdominal cavity are sunken, the back muscles are thin, and the severe ones are blade-shaped. Take gentian as an example, the weight of normal fish is about 50g, while the weight of diseased fish is only 10-20g), and the cumulative mortality rate can reach more than 50% in 4-10 days after stopping feeding, and the serious one is 80-90%. It is popular, and almost all cultured grouper, including pearl gentian, green spot and tiger spot, are infected, and the epidemic area has also expanded to the whole of Hainan and many grouper breeding areas such as Guangxi, Guangdong and Fujian. The farming industry has suffered serious losses

However, due to the small size of the pathogenic agent (a microsporidia), the size of mature spores is 1.3-1.5x1.6-2.4 microns, it is easy to miss detection by ordinary light microscope observation and be confused with other symptoms of grouper white stool. Delayed treatment (such as grouper starvation, feed problems, and bacterial infections can cause white feces to be excreted) confuses

And once mature spores are formed, they are extremely resistant to drugs due to the abnormally hard spore shells. Even if they are diagnosed, they will lose the best period for treatment

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