Multiplex PCR method capable of simultaneously detecting 7 virulence genes of Aeromonas

Abstract

The invention provides a multiplex PCR method capable of simultaneously detecting 7 virulence genes of Aeromonas. The invention comprises the following steps: screening of primer combination and primer sequences; a preparation method of pathogenic bacteria DNA template; multiplex PCR reaction system and PCR amplification; and multiplex PCR product detection and result interpretation. The multiplexPCR detection method can efficiently, specifically and sensitively detect and identify the presence of Aeromonas in a sample and 1-7 virulence genes through two PCR reactions. The method is applicable to the rapid identifying of pathogenic strains and common strains of Aeromonas fish, and is applicable to molecular-assisted diagnosis of fish Aemmonasis and confirmed diagnosis of such disease. Themethod is applicable to the whole process monitoring of aquaculture animal breeding, is applicable to different sample types including water bodies, sediments, farm animals and aquatic products, monitors whether the sample contains Aeromonas or not and that the strain is common or pathogenic Aeromonas, and can identify the virulence gene types and relative expression levels of pathogenic Aeromonas.

Description

technical field [0001] The invention relates to the detection of pathogenic microorganisms in aquatic animals, in particular to a multiplex PCR method capable of simultaneously detecting seven virulence genes of Aeromonas in fish (especially eel) pathogenic bacteria. Background technique [0002] Eel (Angulla spp.), commonly known as eel, is an important economic fish in the world because of its delicious meat and high nutritional value. China's eel farming production accounts for about 70% of the world, with an annual output value of more than 10 billion yuan. In the current high-density intensive culture of eel, aeromonas disease occurs frequently, causing serious harm and huge economic losses. Studies have found that after infection with different strains of the same Aeromonas, the symptoms of eels are very different, including gill rot, red skin, perforation, hemorrhagic sepsis, enteritis and epidemic ulcers; on the contrary, different species of Aeromonas However, eel...

AI Technical Summary

Problems solved by technology

Previous studies have found that aerolysin (Aer), hemolysin (hlyA), enterotoxin (act, ast, alt), serine extracellular protease (ahp), heat-sensitive extracellular protease (epr) (Xiong Jing, Yu Qin , Guo Songlin, et al. Comparison of extracellular products and activities of virulence genes of seven eel pathogenic Aeromonas strains. Huazhong Agricultural University Journal, 2017; 36(1):76-85) and other extracellular toxins are the source of eel The most common major virulence factor of Aeromonas is highly toxic to eels and causes significant economic losses. However, so far, no relevant virulence genes have been detected and monitored in the breeding process

Images